Home

draft - Thermo Fisher Scientific

1. 8 10 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued Item Description Labels Controls how labels are viewed beneath an electropherogram E No Labels v Bins Ctrl Shift B Horizontal Labels Peak Positions Vertical Labels e No Labels Labels are off File Edit View Tools Alleles Help Plot Setting HD Genotyping ZAE Panes fs NT Ma FEE al FE Fi Fl pe PT a AOS es Sample Name Pand sQo SQ Sampled Tanie vi C a D21511 D75820 CSFIPO D8 1179 100 110 120 130 140 150 160 170 180 1390 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 for essary aesszg 100 0o 120 430 440 450 160 170 180 190 200 20 220 230 240 250 260 270 280 290 300 30 320 330 340 350 800 Sample Ideni v1 C DERESE 100 o 120 130 140 150 160 170 180 190 200 20 220 230 240 250 260 270 280 290 300 30 320 330 340 350 Sampled Ideniifler v1 D E 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 Izle KI X 201 71 _Y 420 Bin 13 3 Marker TH01 e Horizontal Labels Labels are positioned horizontally across the screen Labels in close proximity overlap with the left most label displayed on top Click any label to bring it to the top for viewing 5 x EE Edit View Tools Alleles Help Plot Setting
2. SQ Sizing Quality e A value Pass I Check or Low Quality is automatically displayed to indicate the sizing process quality e The sizing quality is calculated based on the similarity between the size standard fragment pattern and the actual size standard peak distribution pattern in the sample e Not editable e This feature performs the following functions Identifies and eliminates the primer peaks based on peak shape Performs size matching ratio matching Makes a size calling curve using the Local Southern Method PQV Flags on the Genotypes Page The PQV flags on the Genotypes page Column Description AE Allele Edit check box e The check box displays unchecked immediately after analysis e The check box is checked when the marker allele calls have been edited by you Note The marker allele calls are edited in the Plot View page The user can enter comments in the Allele History Comments column on the same page A 6 GeneMapper ID Software v3 1 User Guide Using Process Component Based Quality Values The PQV flags on the Genotypes page continued Column Description ADO Allele Display Overflow check box e The check box is checked when the number of alleles the marker calls exceeds the number to display previously set by you e The user specifies how many alleles to display in the Table Settings Editor parameters The default is two e
3. Did you receive any error messages or alerts If so what did they say if possible take screen captures At what point during analysis did these errors alerts occur Is the problem reproducible H 4 GeneMapper ID Software v3 1 User Guide Troubleshooting Sheets Steps taken prior to encountering or to reproduce the problem L Size standard default or custom Peak detector mode classic or advanced Instrument model Dye Set F or G5 Run module GeneMapper ID Software v3 1 User Guide H 5 Appendix H Troubleshooting Sheet H 6 GeneMapper ID Software v3 1 User Guide GeneMapper ID Software Verification This appendix covers Summary of GeneMapper ID Software Verification 2 AmpF STR Kits Samples Instrumentation Hardware and Software Used for Verification Testing 4 Results of Verification Testing naonnana 00 cee ee eee 7 COMCIUSIONSs tect ise ee eed ner at eet aan EEE I 39 RELECO espana E E ne ate We E e aie e e Te dete e ENA I 40 GeneMapper ID Software v3 1 User Guide l 1 Appendix GeneMapper ID Software Verification Summary of GeneMapper ID Software Verification GeneMapper ID Software v3 1 verification was performed to provide documentation to confirm the functionality of the software features The test plan which Applied Biosystems has defined as verification of the software was designed to devel
4. Item Description For example Polynomial Degree Sets the degree of the polynomial Min setting Max setting 2 5 Peak Window Size Sets the width of the window Min setting Max setting 1 above the Degree of Polynomial differentiation setting Number of data points between peaks These parameters control the sensitivity of this process Sensitivity increases with the polynomial degree and decreases with the window size Use polynomials of degree 2 or 3 for well isolated peaks such as those from a size standard and a degree 4 for finer control For degree 4 the Peak Window Size should be 1 to 2 times the full width at half maximum of the peaks that you wish to detect These parameters cannot be set for each color independently Slope Threshold for Peak Start Slope Threshold for Peak End Determines where a peak starts and stops A peak ends when the first derivative again exceeds the Slope Threshold for Peak End Slope Threshold for peak start must be non negative and Slope Threshold for peak end must be nonpositive Values other than O will move the extent of the peak toward its center GeneMapper ID Software v3 1 User Guide Peak Detection Polynomial Degree and Peak Window Size Parameters Peak Detection Polynomial Degree and Peak Window Size Parameters About These Parameters How These Param
5. Size Standard The list is editable via the Size Standard Editor in the Size Standards tab in the GeneMapper Manager window Matrix The list is editable via the Matrix Editor in the Matrices tab in the GeneMapper Manager For the ABI PRISM 377 DNA Sequencer and the ABI PRISM 310 Genetic Analyzer instruments the matrix file is created in the Matrices Tab in the GeneMapper Manager window Run Name From sample sheet not editable GeneMapper ID Software v3 1 User Guide C 5 Appendix C Project Window Software Interface Samples view columns and descriptions continued Column Description Instrument Type From sample sheet not editable Instrument ID Run Date and Time From sample sheet not editable From sample sheet not editable REF SQI Reference data indicates that the selected sample is defined as reference data in the Panel Manager Sizing quality invalidated checkmark indicates that sizing quality value is 1 0 User Defined 1 3 Also from the vertical bar enabled in the sample sheet fields See Chapter 2 Getting Started with GeneMapper ID Software Editable free text Note Refer to Appendix A for PQV information Samples View Features Feature Description Select Row s Click the Row Header box drag to select a continuous range Select Column s Click the Column Header box drag to select a continuo
6. X 269 75 Y 427 Bin 9 2 Marker D75820 8 14 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued Item Description Allele Changes Shows hides label editing effects e Turned Off Deleted label disappears Edited Added label looks normal e Turned On labels manually edited are shown with descriptors and the allele call Label deleted Allele label is shown with a diagonal slash through it BT y TE r Label changed Allele label is shown with a double upper bar d Chagzed alleles Label added Allele label is shown with a double upper bar Le F Eu Controls To Top Controls whether or not the control samples are displayed at the top of the Plot window Checked when active When active a new pane is displayed at the top of the Plot window Only samples and their corresponding genotypes designated as Control or Allelic Ladder in the Sample Type column of the Project window display in this pane If multiple controls are shown use the scroll bar to scroll through the different controls Note Any samples designated as Positive Control Negative Control and or Allelic Ladder in the Sample Type column are affected If only the controls are selected only one panel is displayed with a scroll bar function GeneMapper ID Software v3 1 User Guide Chapter 8 Using Plot Windows Samples an
7. a Select the appropriate user name b Enter your password c Change the database host if necessary d Click OK Note For information on how to create new users and passwords see Users Tab on pages C 24 through C 25 2 8 GeneMapper ID Software v3 1 User Guide Logging Onto and Logging Out of the GeneMapper ID Software Database Host Option The New Host option allows you to connect to GeneMapper D software v3 1 databases on other computers from the Login screen 1 Click the New Host button on the Login Screen to open the New Host dialog box il New Host Enter new GeneMlapper host information Host Marne Machine Type stand alone 2 Inthe Host Name field enter the machine name or IP address of the database host The window displays the new database host and its user names list Note If the GeneMapper D software cannot connect to the database host you entered the following error message opens You have entered an invalid host Click OK to exit and reenter the database host information 3 Select the appropriate machine type from the drop down list 4 Click OK Note Multiple copies of the GeneMapper D software cannot share the same database If you want to have the same project analysis method and size standard definition as another GeneMapper D database you need to use the import and export functions in GeneMapper Manager and Panel Manager IMPORTANT Only one
8. ee l l L WF OD Tae setting fio Tae 100 8 3 Project E gt Casework CO_Control fsa CO Ladder fsa CO_Samplet fsa CO_Sample2 fsa PP_Control fsa H PP_Ladder fsa PP_Samplet fsa PP_Sample2 fsa info Raw Data EPT Data CO_Ladder fsa 0 1000 EP Voltage ME EP Power 2000 3000 4000 5000 6000 EP Currert M RunTemp Line Description Blue EP Voltage Black EP Power Green EP Current Red Run Temperature 3 8 GeneMapper ID Software v3 1 User Guide Overview of the Project Window There are several plot scaling features in the EPT view e Zoom in on one area by placing the cursor in either the X or Y axis label areas and click and drag The software zooms in on the area of selection e Return to a full scale plot by double clicking in the label area again The plot window 1s restored to full scale e Scale the Y axis to maximum Y or a user defined value by selecting the Y Axis Scale from the View menu or by specifying upper and lower limits in the Scale to dialog box Sample Columns Use columns to set up analyses after you have added samples to a project Column Menus The columns in the Samples view of the Project window have drop down menus to apply settings to selected cells in the column e Sample Type Positive control Negative control Allelic ladder
9. Note New profiles can only be created through the GeneMapper Manager Plot Settings Opens the Plot Settings Editor to allow the currently Ctrl T selected Plot Setting profile to be edited Plot Settings Editor xX General Sample Header Genotype Header Sizing Table Labels Display Settings r Sample Header Settings Show Column Sample File 1 2 Sample Name al M Panel 4 M Off scale 5 Sizing Quality Show Hide i OK Cancel 8 22 GeneMapper ID Software v3 1 User Guide Plot Window Menus Alleles Menu The Alleles menu used to edit allele call labels on selected peaks is only enabled when labels are displayed in the Plot window and one or more peaks are selected lel Altes iila Delete Allele Delete History Rename Allele Allele Edit Comment Ctri 0 Add Bin Ctrl B Edit Bin Ctri U Delete Bin Delete Alleles menu items and descriptions Item Description Editing Mode Controls whether the action of the mouse within the electropherogram is in Peak Selection or Binning mode Editing Mode Peak Selection Ctri mod Allele ipkel Binning Ctrl In Peak Selection mode you can e Select peaks e Edit allele calls Clicking on a peak automatically highlights that peak s data row within the Sizing Table or Genotype Table if an allele call has been made e Select peaks continuously Use the Shift ke
10. If previous Project has pending changes the following alert message is displayed you want to save changes Yes No Cancel blank Project is displayed Always enabled Save Project Ctrl S Saves Project to the file named at the start of analysis Named projects are saved to the GeneMapper D database without a dialog box Enabled when the Project table has pending changes Save Project As Displays the Save dialog box Always enabled Add Samples Opens the Add Samples To Always enabled To Project Project dialog box Ctrl I Export Table Displays the Export table dialog Enabled when the Ctrl E box Project table Exports table as tab or comma eomiaiis Gata delimited text file Print Displays the Print dialog box Enabled when the Ctrl P project table The standard Print Setup dialog is opened from the Print dialog contains data GeneMapper ID Software v3 1 User Guide Appendix C Project Window Software Interface Item Description Enabling Log Out Closes the Project window and displays the Login window Displays a Save alert message if the Project has pending changes Always enabled Exit Alt F4 Exits the GeneMapper ID application displays Save alert message if Project has pending changes Always enabled Edit Menu The commands in the Edit menu are used to manage the contents of the Project wind
11. Table Filter a e Show All Rows Ctrl F Header Show Selected Rows Ctrl G Warker Range Ctri R Hide Selected Rows Ctrl H Marker Indicator Ctrl k show Allele Call Rows Ctrl V e Show All Rows Shows all available data rows e Show Selected Rows Shows only peaks that are selected in the electropherogram s e Hide Selected Rows Hides all peaks that are selected in the electropherogram s e Show Allele Call Rows Shows only peaks that have allele calls assigned This control is applicable to the Sizing Table only GeneMapper ID Software v3 1 User Guide Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued ltem Description Header Controls whether the individual Sample or Genotype Header is shown above an electropherogram Checked when active Note The information shown in the Header can be configured through the Plot Setting Editor e Headers Turned On K Samples Plot File Edit View Tools Alleles Help Plot Setting HID Genotyping gt Panes fa a FE al WE Fc af T a SF 8 Pand SQO SQ l01x D75820 CSFIPO D81179 Dzis 10 120 130 440 150 460 170 180 190 200 240 220 230 24 250 260 270 280 290 300 310 320 330 340 350 360 sangis Identititer_v1 C ID3 1358 THOI DI3S317 ID16 539 D21338 ao t 0 10 150 160 A70 180 190 200 210 220 2390 240 250 260 270 200 290 390 810 320 390 340 35
12. 2 S e e a A TT 1 2 3 Column Description 1 Marker name locus name 2 Control allele names as comma separated bin names A G if none there must be a dash 3 Comments if none write none Bins Definitions The examples below show SNP genotyping bin definitions in the for SNP format used by GeneMapper JD software v3 1 in a tab delimited text Genotyping file txt using notepad or wordpad or an Excel spreadsheet saved as a txt file Note ASR Total allele size range The allele size data automatically exports and then the range is printed 4 30 GeneMapper ID Software v3 1 User Guide Formats of Panel and Bin Text Files File Edit Format Help Chemistry Kit SNPdema Binset Name SNPdema Panel Name ho lexdema Marker Name 20mer ASR 24 6 26 1 A 24 6 26 1 Green Marker Name zamer ASR 2 3 29 9 2f 3 29 0 Blue 28 5 29 9 Green Name s6mer 38 4 4 2 38 4 40 1 Blue Name 44imer 46 4 43 4 46 8 48 8 Red Name Sener 53 1 55 2 53 1 54 6 Yellow 54 0 55 2 Red Name 6omer 61 4 63 0 61 4 63 0 Yel low ia 1 2 3 4 Column Description 1 Bin name The name of the allele at an expected location within a locus 2 Bin size range minimum 3 Bin size range maximum 4 Bin color GeneMapper ID Software v3 1 User Guide 4 31 Chapter 4 Using Panel Manager 4 32 E4 Microsoft Excel Book3 TST ES File Edit wiew Insert Format Tools Data Window Help OD c
13. Copies Collate options Check box enables collating of copies GeneMapper ID Software v3 1 User Guide C 33 Appendix C Project Window Software Interface Printing Tables and Electropherograms Introduction All tables and electropherograms can be printed as what you see is what you get WYSIWYG except for these conditions e Only the number of columns shown in the table printout below are printed out on a single sheet of paper in the Portrait mode Additional columns to the right are printed out on additional sheets of paper Change to the Landscape mode to minimize the number of sheets of paper used Note To reduce the number of sheets of paper printed for a table hide columns you do not want to print using the Table Settings Editor e Printed plots are sized vertically to fill a single sheet of paper With four plots the plots are sized close to the same size as the default size on your monitor Six plot panes are a practical limit for a single sheet C 34 GeneMapper ID Software v3 1 User Guide Example Table ra Database_Samples Notepad Printing Tables and Electropherograms O x Printout Bile Edit Format Help Sample Name Run Name Panel Marker Dye Allele 1 Allele 2 a ID_Neg_cntr Databasing Identifiler_ v1 D8S1179 6 ID_Neg_cntr Databasing Identifiler_yvl D21511 6 ID_Neg_cntr Databasing Identifiler_vl D7S820 6B ID_Neg_cntr Databasing Identifiler_vl CSF1P 6 ID_Neg_cntr D
14. Headquarters 850 Lincoln Centre Drive Foster City CA 94404 USA Phone 1 650 638 5800 Toll Free In North America 1 800 345 5224 Fax 1 650 638 5884 Worldwide Sales and Support Applied Biosystems vast distribution and service network composed of highly trained support and applications personnel reaches 150 countries on six continents For sales office locations and technical support please call our local office or refer to our Web site at www appliedbiosystems com www appliedbiosystems com Applied Bisystems Applera Corporation is committed to providing the world s leading technology and information for life scientists Applera Corporation consists of the Applied Biosystems and Celera Genomics businesses Printed in the USA 12 2003 Part Number 4838775 Rev C an Applera business
15. IMPORTANT To properly install GeneMapper ID software v3 1 on the computer that is connected to 3100 3100 Avant system using Data Collection v2 0 the data collection software must be running If data collection is not running the GeneMapper ID software v3 1 does not register with the Data Service software e GeneMapper JD software v3 1 runs on Windows NT and Windows 2000 platforms only Conversion utilities are included for analyzing data from Macintosh based sample files e The version of Oracle database in the GeneMapper JD software v3 1 is an embedded license for use by five 5 named users IMPORTANT To accommodate more than five 5 users additional GeneMapper ID software or Oracle database licenses must be purchased GeneMapper ID Software v3 1 User Guide 2 3 Chapter 2 Getting Started with GeneMapper ID Software e When installing GeneMapper ID software v3 1 you must log into the local machine with administrator privileges e TCP IP has to be installed prior to the installation of the Oracle database Installing To install GeneMapper JD software version 3 1 use the GeneMapper ID nstallationReadMe txt file on the CD ROM as shown in Figure 2 1 Software iz www_appliedbiosystems com MAIN MENU Installation Readme About GeneMapper Software v3 1 Install GeneMapper Software v3 1 Install Adobe Acrobat Reader 5 0 EXTRAS MENU Software Extras GeneMapper ID Soft
16. Sample e Analysis Method e Panel e Size Standard e Matrix e Specimen Category Note To genotype your samples select Allelic Ladder in the Sample Type column in the allelic ladder row Note The Panel column displays a pop up window that contains a hierarchical list of kits and panels A panel is required for any allele calling analysis Use each of these columns to apply needed analysis parameters to selected samples You can make different list selections to the individual column rows representing samples GeneMapper ID Software v3 1 User Guide 3 9 Chapter 3 Using the Project Window Rules for Columns The following rules apply to column selections e Clicking the header for a column selects the entire column e Shiftt clicking selects a continuous selection range e Ctrl clicking individual cells makes discontinuous selections Applying a List Selection to an Entire Column To apply a Menu selection 1 Apply the selection to the top cell in a column a Select the top cell in a column to display a drop down list of items b Select and click an item to apply the selection to the cell 2 Select all other cells in the column either as a continuous or discontinuous selection to which the list selection is to be applied 3 Click Edit gt Fill Down Ctrl D to apply the choice made for the first cell to all selected cells Filling Down a Column To fill down a column 1 Double click the top cell in a co
17. daa a 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 230 290 300 310 320 330 340 350 D351358 2 E 100 110 120 130 140 150 160 170 180 130 200 2410 220 230 240 250 260 270 2380 290 300 310 320 330 340 350 D5s818 100 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 X 234 75 Y 6000 Figure l 4 Female single source DNA Sample amplified with the AmpF STR Identifiler PCR Amplification kit electrophoresed on a 310 Genetic Analyzer for Windows NT OS then analyzed with GeneMapper D software using the Advanced algorithm Genotyping The second parameter evaluated in the Peak Detection and Genotyping category is the genotyping evaluation comparing the results from data analysis with GeneScan Software v3 1 2 and Genotyper software v2 5 2 Macintosh OS to GeneMapper ID Software using the Classic algorithm and GeneScan Software v3 7 1 and Genotyper software v3 7 Windows NT OS to GeneMapper ID Software using the Advanced algorithm The analysis parameter settings for the software packages were defined according to Applied Biosystems recommendations Applied Biosystems 2002 with a GeneMapper ID Software v3 1 User Guide I 13 Appendix GeneMapper ID Software Verification defined peak amplitude threshold PAT of 50 RFU Five observations were documented when comparing the results from GeneScan and Genotyper software to the results of Gen
18. 5 62 Matrix Result Oo aU i D B G v R 0 1 0000 0 2009 0 1176 0 0319 0 0385 0 2897 0 3527 0 7938 1 0000 1 175 0 0404 0 0962 0 1992 0 304 1 0000 J Cancel For each dye the value where the dye fluorescence is read by the appropriate filter is 1 000 The adjacent colors show the amount of overlap for which the system must compensate The adjacent values in most cases should be less than 1 000 but greater than or equal to 0 0000 GeneMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix Assigning the Matrix to Samples Introduction After generating the new matrix assign it to all the samples that you want to analyze IMPORTANT After assigning your matrix to samples refer to Appendix C Project Window Software Interface for information on evaluating the matrix Procedure To assign a matrix to samples l Open or create a GeneMapper ID Project containing the samples you wish to analyze Select the Matrix column in the project window Select the appropriate matrix from the drop down list for each sample or use the Edit gt Fill Down feature Apply additional project settings prior to analyzing the sample files Using the Matrix with Data Collection Using Different Be sure to copy matrices generated and exported on the analysis Computers for computer the computer running GeneMapper ID software to the Data Collection Data Collection computer if di
19. Always GeneMapper D window and displays the software enabled version C 26 GeneMapper ID Software v3 1 User Guide Project Window File Menu Dialog Boxes Project Window File Menu Dialog Boxes Introduction The following dialog boxes are accessed from the Project window File menu This section describes each of the dialog boxes Note This section contains information on all application dialog boxes except for the Add Samples to Project dialog box which is covered under its own section See Add Samples to Project Dialog on page C 37 Save Dialog Box The Save dialog box accessed from the New Project window is presented when a project already exists in the Project window and provides you with the option of saving the project A Save Project D oe G Project name on GeneMapper ID Software v3 1 User Guide C 27 Appendix C Project Window Software Interface C 28 Open Project Dialog Box The Open Project dialog box displays the list of projects in the database You use the first field to find a project quickly then click the Open button to open the selected project or double click the project name r IF Open Project 1 Type the first few letters of the project you are looking for 2 Select the project you want then click Open mma aio of Samples E Casework Project 2002 06 05 eres g 3 Casework Project 06 2002 06 05 15 22 29 8 4 HID Case
20. Analysis Completed stop Samples view columns and descriptions Column Description Sample or row From application used to select a single entire row number C 4 GeneMapper ID Software v3 1 User Guide Overview of the Project Window Samples view columns and descriptions continued Column Description Status A La symbol indicates that the sample needs to be analyzed or reanalyzed Note These symbols indicate status of each sample and they control which samples will be analyzed during the next analysis if the Analyze icon is selected There are other analysis options which ignore the status field e The Status field next to each sample contains the La symbol prior to analysis This field is empty after analysis if analysis was successful e Resetting parameters such as the Panel Size Standard Sample Type and the Analysis Method resets the symbol for a given sample enabling reanalysis Sample File From sample sheet not editable Sample Name From sample sheet editable free text Comments Free text Editable Sample Type Valid types are Sample Control Allelic Ladder and Primer Focus Negative Control Editable via Sample Type drop down list box Analysis Method The list is editable via the Analysis Method Editor in the Analysis Methods tab in the GeneMapper Manager window Panel Panel window contents come from Panel Manager Editable via Panel window
21. Analysis Type HIL OK Cancel Figure 5 10 General Tab of the Analysis Method Editor Note The Analysis Type reflects the choice selected in the New Analysis Method dialog box GeneMapper ID Software v3 1 User Guide 5 15 Chapter 5 Using GeneMapper Manager Allele Tab In the Allele tab you can set a number of parameters controlling allele calling including e Bin Set e Marker Repeat Type for microsatellites e Amelogenin Cutoff value e Range Filter Note Applied Biosystems recommends that you leave all other parameters in the default setting after selecting the AmpF STR bin set for data analysis Analysis Method Editor HID x AmpFISTR Bins w1 7 Figure 5 11 Allele Tab of the Analysis Method Editor GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab Bin Set Bins are a way of matching your data to the allele definitions contained in the panel Bins are the locations in which you expect an allele to size Alleles are defined in bins centered around the average size in base pairs for example 110 00 0 5 bp Sample peaks that fall within a bin are genotyped accordingly You must choose a bin set for your analysis in order to guide the algorithms in assigning allele calls This drop down list enables you to choose the bin set to be used for analysis The bin set chosen for analysis must match the kit panel chosen for analysis Note For AmpF STR analysis you import the bi
22. Chapter 4 Using Panel Manager Column Descriptions 1 Marker name locus name 2 Dye color 3 Minimum ASR allele size range actually marker size range 4 Maximum ASR allele size range actually marker size range 5 Control allele names as comma separated bin names 1 2 Note If none there must be a dash 6 Type of repeat 2 dinucleotide 3 trinucleotide 4 tetranucleotide 5 non repeat 7 Marker specific stutter ratio if none use zero 0 0 8 Comments if none write none 9 Alleles present in the allelic ladder for each marker 4 24 GeneMapper ID Software v3 1 User Guide Formats of Panel and Bin Text Files i AmpFLSTR_Panels_v1 Notepad File Edit Format Help PGenemMapper ID v3 1 Last edited 071403 a GMv3 0 for import into GeneMapper ID v3 1 Version GMy3 0 Kit type Microsatellite Chemistry kit AmpFLSTR_Panels_v1 Panel Blue_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 VWA blue 151 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none L8 19 20 21 22 23 24 25 26 26 2 27 28 29 30 Panel Green_I_v1 AMEL green 106 00 114 00 x 9 0 060 none os ie THOL green 159 00 205 00 8 393 3 4 0 060 none 5 6 7 8 9 9 3 10 TPOX green 212 00 254 00 8 4 0 060 none 6 7 8 9 10 11 12 13 CSFLPO green 275 00 323 00 10 12 4 0 090 none Gp 7p8 9 10 11 12 13 14 15 Panel Profi
23. Ctrl Shift N Y Axis Scale The Y Axis Scale option is only enabled when viewing the raw data or EPT data of a sample file Y Axis Scale Scale to maximum Y show Navigator Ctrl Shift hi Scale to There are two options e Scale to maximum Y scales to the maximum Y value for the selected sample e Scale to opens a dialog box in which a user can define the value of the Y axis GeneMapper ID Software v3 1 User Guide F Enter Axis Scale Yalues E x Minimu p aua Maimun s000 Cancel Project Window Menus Tools Menu The Tools menu provides access to the GeneMapper Manager Panel Manager and Table Setting Editor applications stim Help GeneMapper Manager Cirl h Panel Manager Table Setting Editor Ctri T Cirl J show Offsets Options Item Description Enabling GeneMapper Opens the GeneMapper D Always enabled Manager Ctrl M dialog box See Chapter 5 Using GeneMapper Manager Panel Manager Ctrl J Opens the Panel Manager dialog box See Chapter 4 Using Panel Manager Always enabled Table Setting Editor Ctrl T Opens the Table Setting Editor dialog box See Table Settings Tab on page 5 27 samples are present Only Table Profile Only enabled when allows editing of a currently selected GeneMapper ID Software v3 1 User Guide Appendix C Project Window Software Interfa
24. GeneMapper ID Software Version 3 1 Human Identification Analysis User Guide Applied Biosystems Copyright 2003 Applied Biosystems All rights reserved For Research Forensic or Paternity Use Only Not for use in diagnostic procedures Notice to Purchaser License Disclaimer Purchase of this software product alone does not imply any license under any process instrument or other apparatus system composition reagent or kit rights under patent claims owned or otherwise controlled by Applera Corporation either expressly or by estoppel GeneMapper ID Software version 3 1 has undergone a verification process defined by Applied Biosystems However human identification laboratories analyzing forensic paternity databasing and single source samples that choose to use GeneMapper ID Software for data analysis should perform their own appropriate validation studies Information in this document is subject to change without notice Applied Biosystems assumes no responsibility for any errors that may appear in this document This document is believed to be complete and accurate at the time of publication In no event shall Applied Biosystems be liable for incidental special multiple or consequential damages in connection with or arising from the use of this document Purchase or receipt of this software does not include or guarantee any training by persons belonging to Applied Biosystems or Applera Corporation ABI PRISM AmpF
25. GeneMapper ID Software v3 1 User Guide Project Window Edit Menu Dialog Boxes Project Window Edit Menu Dialog Boxes Find Dialog Box The elements of the Find dialog box is described below Find Ea Find what in column Sample File Name x Element Description Find what Field containing the search string In column List box containing the names of columns in the table Find Next Finds the next instance of the search string in the specified column and selects it Find All Finds and selects all rows in the table that contain the search string in the specified column Close Closes the Find dialog box Table selections remain intact GeneMapper ID Software v3 1 User Guide C 45 Appendix C Project Window Software Interface Sort Dialog Box The Sort dialog box shown below allows sorting of sample files and results by ascending or descending order in up to three steps Sort by Ascending Sample File T p U Descending Then by Ascending Sample File Y a O Descending Then hy Ascending Sample File T T gt Descending ok Cancer The Sort dialog box has three entry fields for entering sort criteria the Then by fields allow successive sorting by two other criteria Element Description Sort by Sort box containing the column which will be sorted Then by Ascending Option buttons for choosing if th
26. ID Software v3 1 User Guide 9 5 Chapter 9 Exporting Table Data CODIS Export Manager The CODIS Export Manager is used to view and set values for three fields required for exporting samples to CODIS e Specimen Types e Source Lab ID e Destination Lab ID Setting CODIS CODIS laboratories should use their assigned Source Lab ID and Export Fields Destination Lab ID To set CODIS export fields 1 Select Tools gt CODIS Export Manager Alleged Father Alleged Mother Biological Child gt fo export Add Delete Source Lab IDs srciab Add Delete Destination Lab IDs Jdestiab Add Delete OK Cancel 9 6 GeneMapper ID Software v3 1 User Guide CODIS Export Manager 2 View the Specimen Types Specimen Types Biological Child gt Y fo export Add Delete The specimen types included in GeneMapper ID Software version 3 1 are currently accepted by CODIS When CODIS accepts a new specimen type you can add the specimen type to the software by typing it in the text box and then clicking Add 3 Adda Source Lab ID Source Lab IDs a Enter your assigned source lab ID in the text box Note The source lab ID cannot exceed nine characters b Click Add 4 Add a Destination Lab ID Destination Lab IDs destiab Add Delete a Enter your assigned destination lab ID in the text box Note The destination lab ID cannot exceed nine characters b Click
27. Set Size Standard to c ee F Ho _ss500 for all samples Read fromthe Sample Set 310377 Matrix to gt but for all samples Read fromthe Sample Select a Panel for all samples Read from Data collection Comment Panel field CPBample z for all samples Read from Data collection Info field OK Cancel Element Description Set Analysis Method to When you add samples the Analysis drop down menu and Method property is set to the value in the text box which contains the list of analysis Read from the Sample methods from the GeneMapper D option button database Set Size Standard to When you add samples set the size drop down menu and standard property to the value in the text box which contains the list of size standards from the GeneMapper D database Read from the Sample option button GeneMapper ID Software v3 1 User Guide 6 5 Chapter 6 Options Dialog Box 6 6 Element Description Set 310 377 Matrix to drop down menu Read from the Sample option button Set the matrix file to the dye you are using or read from the sample file Set Panel to e Select a Panel for all samples option button and window e Read from the Data collection Comment Panel field option button When you add samples the panel for all samples is set to the value in the text box When the option button is selected the following
28. e E sizing table as shown in the Samples Plot view Figure 5 26 Sizing Table Tab of the Plot Settings Editor The Show button displays all selected rows as visible checked and the Hide button displays all selected rows as hidden unchecked GeneMapper ID Software v3 1 User Guide Section 5 4 Plot Settings Tab Labels Tab The Labels tab controls how labels are viewed for each allele You can display up to four labels per peak Plot Settings Editor General Sample Header Genotype Header Sizing Table Labels Display Settings Show Labels When opening the Plot Windows Label 1 Allele Call fy Show data type prefixes Label 2 ize W Show type of edit Label 3 W Invert mutant labels Label 4 None Label Color Dye Color Border Font Times Mew Roman Size 10 E Figure 5 27 Labels Tab of the Plot Settings Editor Use the drop down menus to display the labels you want to use Plot Settings Editor General Sample Header Genotype Header Sizing Table Labels Display Settings Show Labels When opening the Plot Window Label 2 Size Show type of edit Label 3 Area T W Invert mutant labels Label 4 Mane Label Color Dye Color Border Font Size GeneMapper ID Software v3 1 User Guide 5 47 Chapter 5 Using GeneMapper Manager Table 5 13 Labels tab menu items Name Type Description Allele Call Area Drop down menu Labels determined by H
29. Baseline Window 51 pts Size Calling Method 2nd Order Least Squares 3rd Order Least Squares Cubic Spline Interpolation Local Southern Method C Global Southern Method Peak Window Size Slope Threshold Peak Start Peak End Factory Detautts OK Cancel Figure 5 8 Analysis Method Editor GeneMapper ID Software v3 1 User Guide 5 13 Chapter 5 Using GeneMapper Manager New Analysis When you create a new analysis method the following dialog box Method Dialog opens prior to the analysis method editor This dialog box allows you Box _ to select the analysis type for your samples and controls the items available for customization within the Analysis Method Editor tab Hew Analysis Method Select analysis type HIC C SNaPshot Microsatellite OK Cancel Figure 5 9 New Analysis Method Dialog Box Note Select HID for AmpF STR data analysis 5 14 GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab General Tab In the General tab enter a name and description for the analysis method Create an analysis method description for your specific platform i e capillary or gel based electrophoresis Capillary and gel based electrophoresis may require different start and stop points in the analysis range or peak amplitude thresholds Analysis Method Editor HID E x Analysis Method Description Mame HID Advanced Description Instrument
30. D projects are exported Note Only one preset file extension ser can be used 7 Cancel button Closes the Export Project window without exporting a project GeneMapper ID Software v3 1 User Guide Section 5 1 Projects Tab Import Project The Import Project window enables you to import project data Window previously saved See an explanation of each area in Using the Import Project Window on page 5 8 N F Import Project Look in GeneMapper 5 File nare import Files of type Java serialized file ser Cancel Figure 5 5 Import Project Window GeneMapper ID Software v3 1 User Guide 5 7 Chapter 5 Using GeneMapper Manager Using the Import The elements called out in the figure in Import Project Window on Project Window page 5 7 are described in the table below Import Project Callouts Description Click these icons to Move display up one level in main pane Move display to Home level in main pane This is usually Profiles lt user gt e Create a new folder at the present directory level se Present a list of the contents of the EE selected folder Present details of the selected folder Select drive letter and or folder The contents of the folder selected in the toolbar are displayed here Imports the selected project Enter the name of selected project being imported Th
31. F GeneMapper ID 3 1 Database Project gmid Is Logged In RIE Edi Arelysis View TOOS Help AE NLE Table Setting iz Table v H e 5 a Overview of the Project Window Project Window Views Samples View When you launch the GeneMapper JD software the Samples View is the first view you see Use this view to open existing projects import sample files select analysis settings analyze data and view sample PQVs and sizing quality The Samples tab 1s always enabled find it by clicking once on the Project root node in the left navigation pane or by selecting Samples under the View menu GeneMapper ID 3 1 Database Project gmid Is Logged In Ale Edi Analysis wiew foci Help eS SB a BB tente seting Ho Tae Project Samples Genotypes Status Sample F Sample N Sarnple IC Comment Sample T Specimer Analysis Panel Size Star Ma 1 ID_Contre ID_Contre None no expor HID_Adv Identifiler CE_G5_F 2 ID_Ladde ID_Ladde None Allelic La no expor HID_ Adv Identifiler CE_G5_F 3 ID_Neg_c ID_Neg_ None Negative no expor HID_Ady Identifiler CE_G5_F 4 ID_Sarmpl Sarmple3 None Sample no expor HID_Advy Identifiler CE_G5_F 5 ID_Sampl Sample4 None Sample no expor HID_Adv Identifiler CE_
32. STR AmpF STR COfiler Applied Biosystems Genotyper Identifiler LIZ Profiler Profiler Plus SEfiler SGM Plus and SNaPshot are registered trademarks and AB Design AmpF STR Blue AmpF STR Green Applera GeneMapper GeneScan POP 6 Primer Focus ROX and TAMRA are trademarks of Applera Corporation or its subsidiaries in the U S and or certain other countries AppleScript and Macintosh are registered trademarks of Apple Computer Inc This product includes software developed by the Apache Software Foundation http www apache org Copyright 1999 2000 The Apache Software Foundation All rights reserved This product includes software developed by the ExoLab Project http www exolab org Copyright 2000 Intalio Inc All rights reserved JNI Registry is Copyright 1997 Timothy Gerard Endres ICE Engineering Inc http www trustice com Microsoft Windows NT and Microsoft Excel are registered trademarks of Microsoft Corporation Oracle is a registered trademark of Oracle Corporation All other trademarks are the sole property of their respective owners Applera Corporation is committed to providing the world s leading technology and information for life scientists Applera Corporation consists of the Applied Biosystems and Celera Genomics businesses Printed in the USA 12 2003 4338775 Rev C Contents Preface Chapter 1 Chapter 2 Chapter 3 Chapter 4 How to Use This Guide 0 0c ee ees
33. Select And GeneMapper ID software displays the Analysis Tools gt Analysis Methods page Methods GeneMapper Manager _ Then select the F GeneMapper Manager Analysis Methods Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards tab Name Last Saved Owner Instrument Analysis Type Description Default 2003 06 12 13 57 41 Microsatellite HID Advanced 2003 06 12 14 09 41 gtd TT analyzer HID HID_Classic 2003 06 12 14 07 3 gria jarr analyzer HID a Hew HET Sawe Me Import Expari Pelete Panel Tools gt Panel Panel Manager window Manager Manager Alternative Click the icon File Edit Bins wiew BF A AmpFLSTR_Panels_ Blue y Green Ivi Frotiler w1 Profiler _Plus wi cotiler_v1 SGM _Plus_ v1 identifier v4 SE tiler_v1 _ Protiler Plus Cor cotiler Cops w identifier CODIS k a FX ee ee E E Panel Manager Bin Set ampFLSTR Bins_v1 Panel Hame Profiler Pius vi none Cancel GeneMapper ID Software v3 1 User Guide 3 15 Chapter 3 Using the Project Window To see the Select And GeneMapper ID software displays the Size Tools gt Currently Defined Size Standards Standards eee mage te see ne Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Size Standards roje na ysis a a E ngs mgs ric
34. The Criteria subcommand functions in the GeneMapper D Database tab but not in the Files tab and is used to clear all search criteria from the Criteria Selector List see the figure on page C 42 If undo is not possible the following alert message is displayed Clear all items You cannot undo this action Yes No Cancel Enabled when the field is present and an item exists in that field Options Opens the Options dialog box in the Add Samples tab Always enabled GeneMapper ID Software v3 1 User Guide C 39 Appendix C Project Window Software Interface View Menu The View menu of the Add Samples window is used to switch between the three sources for samples and refresh the file directory The commands are described in the table below Files Ctrl 1 GM Database Cirl 2 SEO UE TCE olertia Wass Item Description Enabling Files Opens the Files tab Always enabled Ctrl 1 GM Opens the Always enabled Database GeneMapper D Ctrl 2 Database tab C 40 GeneMapper ID Software v3 1 User Guide Add Samples to Project Dialog Files Tab The Files tab enables you to choose sample files from local or mapped networked storage devices Files pane F Add Samples to Project Edit view Files Gh Database Add To List gt Options Samples To Add pane Samples To Add Fl Casework oi CO_Cortrol tsa oi CO_Ladder tsa i CO_Sam
35. When alleles shown in the Genotypes table are edited in the Plot window the allele call in the table also changes and confidence value indicators in the table turn to grey triangles Size 1 2 etc Peak size for Allele Not editable Height 1 2 etc Peak height for Allele Not editable Peak Area 1 amp 2 Peak area for Allele Not editable Mutation Indicates allele falling within a mutant bin AE Comment Displays last user edited comment C 8 GeneMapper ID Software v3 1 User Guide Overview of the Project Window Genotypes view columns and descriptions continued Column Description ADO Allele Display e The box is labeled with an 7 when the Overflow number of alleles the marker calls exceeds the number to display previously set by you e The user specifies how many alleles to display in the Table Settings parameters The default is two e There are six columns for each allele to indicate name size height area mutation and comment AE Allele Edit e The box displays unlabeled immediately after analysis e The box is labeled with an x when the marker allele calls have been edited by you e The marker allele calls can be edited in the Plot window The user can enter comments in the Allele Edit column while editing allele calls User Defined 1 3 Editable free text Also from vertical bar enabled in sample sheet fields See Chapter 2 G
36. Y Axis Scale t Scale to marker size range Dyes scale to The X Axis Scale menu command controls the horizontal scale of the reference data electropherogram being viewed in the Panel Manager as follows e Scale to maximum The electropherogram being viewed is scaled to its maximum base pair length as collected from the instrument e Scale to marker size range The electropherogram being viewed is scaled to the size range available for the selected marker e Scale to You can define the horizontal range being viewed GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands To zoom in to a specific region on the X axis of the electropherogram e Click and drag with the left mouse button the desired region on the X axis or e Right click on the X axis to open the X Axis Scale menu command options Y Axis Scale axis Scale Scale to maximum 1 I H Dyes Scale to The Y Axis Scale menu command controls the vertical scale of the reference data electropherogram being viewed as follows e Scale to maximum The electropherogram being viewed is scaled to its maximum height as collected from the instrument e Scale to You can define the vertical range being viewed To zoom in to a specific region on the Y axis of the electropherogram e Click and drag with the left mouse button the desired region on the Y axis or e Right click the Y axis to open the Y Axis Scale men
37. and makes allele calls using defined panels and bin sets with its automated genotyping capabilities Leibelt et al 2003 Further a comprehensive concordance study used forensic simulated samples generated with seven different AmpF STR kits The data were processed on six instrument computer platforms ABI PRISM 310 Genetic Analyzer for Apple Macintosh and Microsoft Windows NT operating system OS ABI PRISM 377 DNA Sequencer for Macintosh and Windows NT OS ABI PRISM 3100 l 2 GeneMapper ID Software v3 1 User Guide Summary of GeneMapper ID Software Verification Genetic Analyzer and ABI PRISM 3100 Avant Genetic Analyzer These data were analyzed with GeneScan Software v3 7 1 and Genotyper Software v3 7 Windows NT OS and GeneScan Software v3 1 2 and Genotyper Software v2 5 2 Macintosh OS Boland et al 2003 and these data have been used for the verification of GeneMapper JD Software v3 1 Peak Detection GeneMapper JD software includes three peak detector algorithms or Algorithms modes allowing different levels of user control over data analysis Classic Mode The Classic mode provides the user with the same analysis parameters and produces results similar to those analyzed with GeneScan Software v3 1 2 Macintosh OS This algorithm aids the user in the adoption of GeneMapper JD software for laboratories that currently use the Macintosh OS and want to maintain current analysis parameter settings Advan
38. configuration requirement J 2 technical support for altered configuration J 2 Conventions bold text 1x IMPORTANTS x in this guide 1x italic text 1x menu commands 1x Notes x user attention words x Cubic Spline Interpolation method B 31 Customer feedback on Applied Biosystems documents xi Cut off Value 5 18 cutoff value 7 3 D Destination Lab ID adding 9 7 selecting 9 9 Documentation feedback xi related to this guide x Dye lanes window example of 3 14 E EPT tab 3 8 Example Plot printout C 36 Example Table printout C 35 export file type for CODIS 9 9 Export Project window 5 5 Export Table dialog box C 31 Index 2 F Filter 8 19 Filtering controls 5 35 Find command planning for use of G 4 Find dialog box C 45 G GeneMapper access rules 2 7 accessing subordinate application windows 3 15 logging in and out 2 7 major steps in using 1 5 overview 1 2 registering 2 6 GeneMapper ID Software verification I 2 AmpFISTR kits used l 4 analysis software I 6 conclusion I 39 instrument platform and computer operating system l 6 references 1 40 results I 7 sample types used I 4 GeneMapper Manager Analysis Method Editor 5 12 Analysis Methods tab 5 9 New Analysis Method dialog box 5 14 GeneMapper Software hardware and software requirements 2 2 installing and registering 2 3 required steps 1 5 GeneScan Analysis Sample Sheet importing data from G 2 Genotypes View filter setting 5 37 genotyping alg
39. quality If the matching quality is not so good could be caused by various reasons like high noise background significant ladder allele imbalance or mobility shifts causing significant local nonlinearity the GQ will be yellow even when all the modulators all other flags in Issue are green The Genotype Quality can be displayed as a numerical value instead of a colored symbol Select this preference in the Analysis Tab within the Options window under the Tools Menu Refer to Chapter 6 GeneMapper ID Software v3 1 User Guide 7 31 Chapter 7 Analyzing and Interpreting Data Concordance Checks 7 32 The software is capable of performing three concordance checks e Checking control alleles against expected alleles that are predefined in the panel used e Comparing genotypes for overlapping loci within different AmpF STR kits For example D3S1358 is a locus used in both Profiler Plus and Cofiler Ifthe same sample was amplified with both kits the software can check that both amplifications produce the same genotype for the overlapping locus e Comparing genotypes for overlapping loci within the same AmpF STR kit e g checking the concordance between duplicate amplifications and injections of the same sample This section focuses on the second type of concordance check listed above Requirements for performing a concordance check include e The genotypes must be saved in the local GeneMapper database Thus the sam
40. quality value determination B 2 B 5 size matching B 2 size matching size calling B 2 Allele tab 5 16 allelic bin definitions 7 2 offsets 7 2 allelic ladder analysis method for 7 2 sample type 7 2 Analysis Method Editor Alleles tab 5 16 General tab 5 15 Peak Detector tab 5 19 Peak Quality tab 5 22 Quality Flags tab 5 24 analysis method for allelic ladders 7 2 analysis methods 5 11 analysis parameter settings analysis range options B 12 data processing options B 13 GeneMapper ID Software v3 1 User Guide Applied Biosystems contacting xi customer feedback on documentation xi Services and Support xi Technical Communications xi Technical Support xi Apply button 4 3 4 8 7 15 7 18 Apply To checkbox 8 10 assign specimen type for CODIS 9 8 Assumptions for using this guide ix B backing up data F 7 Bin Set alleles tab 5 16 choosinga 5 16 Bin view cursor location information 4 41 description 4 40 genotype quality symbols 4 41 Marker Allele Size Range ASR 4 41 purpose of 4 39 X and Y axes 4 41 Bins definitions for SNaPshot 4 30 format of text file 4 27 Bold text when to use 1x C Cleaning F 7 cleaning the database F 7 CMF 3 0 file type 9 9 CODIS assign specimen type 9 8 export manager 9 6 modifying columns 9 8 requirements 9 5 Index 1 CODIS continued setting CODIS export fields 9 6 specimen number 9 8 table export 9 8 Web site 9 5 Command activation three ways of C 10 Computer
41. selected it to be displayed in your sample plots highlight samples within the genotype tab and go to Analysis gt Display Plots The PQVs are an easy way to scan through the data and determine which markers loci are problematic without going through the electropherograms for every sample file PQVs are very useful for processing databasing samples or for high throughput analysis Refer to Appendix A for a detailed description of each PQV To use the Genotype PQVs to evaluate your data 1 Within the Genotypes tab select Analysis gt Low Quality to the Top to bring all samples with low quality Red and questionable yellow Genotype Qualities 2 Highlight all the rows with low quality and questionable Genotype Qualities Note Green Genotype Qualities may not need any further assessment Green indicates a high confidence level that the allele calls are accurate 3 Select Analysis gt Display Plots 4 Scroll through the plots using the PQVs to help determine where the problem lies 5 Decide if you agree or disagree with the software e Ifyou agree then you might remove the sample from the project or remove the allele calls for that particular marker e After reviewing a flagged PQV you can override the GQ by right clicking the red or yellow GQ flag You will be asked if you want to override the Genotype Quality of that marker Click Yes All genotype PQVsturn grey and the GQ turns green GeneMapper ID Softwar
42. 075795E 01 J z 1 5455956 05 R 2 1 000 Size Calling Curve Local Southern Method I I I I I I I I I 1200 1400 1600 1800 2000 2200 2400 2600 26800 Data Point Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper I D software v3 1 The Equation L c m m0 LO The equation attempts to describe the reciprocal relationship between the mobility m and the length LO of the standard fragments B 32 GeneMapper ID Software v3 1 User Guide Size Calling Methods for Classic and Advanced Modes How This Method Works This method which is similar to the Cubic Spline method uses the four fragments closest in size to the unknown fragment to determine a best fit line value Only the region of the size ladder near the fragment of unknown length is analyzed Note Size estimates may be off if any of the standard fragments run anomalously The following table lists how the Local Southern method works 1 The fitting constants of the curve are calculated for each group of three neighboring points on the standard A separate curve is created for each set of three points 2 Acurve is then created by using three standard points two points below and one point above the fragment and a fragment size is determined 3 Another curve is created by looking at an additional set of three points one point below and two points above the fr
43. 1 User Guide Peak Detection Options Advanced Mode Peak Detection Algorithm Settings The Peak Detection options locate peaks at the positive to negative zero crossings of the first derivative of the baselined electropherogram The peak detector computes the first derivative at a data point i by fitting a polynomial to a window centered on i Item Description For example Peak Amplitude GeneMapper ID If you leave the default Thresholds software reports to you value of 50 peaks with only those peaks amplitude above 50 whose heights are at are analyzed and least the Peak display in the tabular Amplitude Threshold data OPACO E Lower amplitude peaks Set the dye amplitude still display in the threshold at a levelthat electropherogram but allows the software to are not analyzed and detect peaks but do not display in the eliminate noise tabular data Full Width Half Defines what If this number is large Maximum constitutes a peak the software ignores Full width Use to specify the smallest full width at half maximum for peak detection The range is 2 99 A typical number might be 3 for microsatellites or 10 for SSCPs noise spikes If the peaks in the data are narrow set the value to a low number Experiment with this value to determine the best number for the data GeneMapper ID Software v3 1 User Guide Appendix B Software Genotyping Algorithms
44. 2 GeneMapper ID Software v3 1 User Guide GeneScan 120 Size Standard GeneScan 120 Size Standard About This Size Standard How It Is Prepared GeneScan 120 Molecular Lengths Electrophero gram of GeneScan 120 The GeneScan 120 Size Standard is useful for sizing fragments between 15 and 120 base pairs It is used when analyzing SNaPshot kit samples All aspects of the preparation of the GeneScan 120 LIZ size standard are proprietary Each fragment contains a single LIZ fluorophore The following table lists the GeneScan 120 LIZ denatured molecular fragment lengths nucleotides for the nine fragments Fragment Fragment Length Length 15 62 20 80 25 110 35 120 50 The following screen shows an electropherogram of GeneScan 120 LIZ run under denaturing conditions F Size Match Editor Edit View Size Match Quality 1 0 2456 2rd 2904 3060 3216 3372 3528 3624 GeneMapper ID Software v3 1 User Guide D 3 Appendix D GeneScan Size Standards GeneScan 350 Size Standard About This Size Standard How It Is Prepared GeneScan 350 Molecular Lengths Electrophero gram of GeneScan 350 The GeneScan 350 Size Standard is useful for sizing fragments between 35 and 350 base pairs The native fragments are uniformly spaced to provide accurate size calling The GeneScan 350 Size Standard is prepared by PstI digestion of plasmid DNA followed by l
45. 6 GeneMapper ID Software v3 1 User Guide K 1 Appendix K Workflow for Storing Sample Files Projects and Second Reads 5 Select File gt Open Project Select the project to open then click Open 6 If you are using a different computer the second analyst must a Import the project created by the first lead analyst in the Project tab of GeneMapper Manager b Select File gt Open Project Select the project to open then click Open c Import the same samples Two folders display in the left navigation pane differentiating each set of analyses GeneMapper ID v3 1 Gasework gmid Is Logged In JE x File Edit Analysis View Tools Help fer lar eons bill LU ED alll Be Tate setina beta AEE NE EEE H E Casework Sample Mame Sample Type Specimen Cater Analysis Method Panel sees Standart H E casework 1 CO_Control Postive Contal hoexpot F_Advanced cofier vt CE 2 CO_Ladder Allelic Ladder no export F_Advanced Goie EF F_HID Gd ja Samplet Sampe no exp export oto F_Adh Advanced anced EOfler s1 1 cE F_HID GS a Sample2 Sampe ncexport F_Advanced COfler v1 CE_F_HID_GS B P2 Cortrol Postivec Control no export F_Advanced Protiler_P Plus v1 b E F HID Gd e PF Ladder AlelicLadder no export F_Advanced Profiler Plus v1 F_HD_G5 E oe e e e O O 2 a Sample2 Sample no export F_Advanced Profiler Plus _ v1 Plus vi CE_F F HID G G4 9 CO Control Sampl
46. 6 Classic Mode Peak Detection Algorithm Settings B 7 Advanced Mode Peak Detection Algorithm Settings B 12 Peak Detection Polynomial Degree and Peak Window Size Parameters 0 00 cece eee B 17 Optimizing Peak Detection Sensitivity Example 1 B 20 Optimizing Peak Detection Sensitivity Example 2 B 22 Optimizing Peak Detection Sensitivity Example3 B 24 Peak Detection Slope Threshold for Peak Start and Slope Threshold for Peak End Parameters B 25 Slope Threshold Examples 000 0000 cece ceeees B 27 Size Calling Methods for Classic and Advanced Modes B 29 GeneMapper ID Software v3 1 User Guide B 1 Appendix B Software Genotyping Algorithms GeneMapper D Software Genotyping Algorithms B 2 Overview Five algorithms used in the GeneMapper JD software are discussed Peak Detection Algorithm Size Matching Size Calling Algorithm in this appendix e Peak Detection uses Basic Advanced or Classic mode to detect peaks and process data e Size matching calling matches found peaks to size standards e Binning determines bin centers for genotyping e Allele calling produces a consensus call based on several allele calling algorithms e Quality value determination assigns a quality value to size and allele calling algorithms See Appendix A Process Quality Values for more information This algorithm uses
47. 96 lane upgrade system with the sequencer instrument Multicomponent This process of eliminating spectral overlap is called Definition multicomponenting Applying a matrix to raw data allows you to generate multicomponented data 5 54 GeneMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix Why Is a Matrix A matrix is necessary because the four or five dyes used to label the Necessary fragments fluoresce at different wavelengths and may have spectral overlaps S FAM JOE 60 60 40 0 b Normalized Fluorescence Intensity p 520 340 560 580 600 620 640 Wavelength nm Assigning a Normally the matrix is assigned to a sample file automatically upon Matrix to a generation during or after a run Additionally a matrix can be Sample manually assigned to a ABI PRISM 377 DNA Sequencer sample file or a ABI PRISM 310 Genetic Analyzer sample file from within the GeneMapper JD software When to Assign a__ Before you can successfully analyze ABI PRISM 377 DNA Sequencer Matrix sample files or ABI PRISM 310 Genetic Analyzer sample files using the GeneMapper ID software you must make a new matrix or assign an existing one to a set of sample files Limitations to The matrix is instrument specific You cannot apply a matrix you Matrices made onthe ABI PRISM 377 DNA Sequencer to data you collected on an ABI PRISM 310 Genetic Analyzer nor can you apply a matrix made on one ABI PRIS
48. Add 5 Click OK to save the changes and close the CODIS Export Manager GeneMapper ID Software v3 1 User Guide Chapter 9 Exporting Table Data CODIS Table Export CODIS Specimen The CODIS Specimen Number up to 24 characters is accessed Number from e The Sample Name field if the UD1 User Defined 1 column is not used e The UDI column if the UD1 column is used Assigning Before you can export a table in the CODIS format a specimen type specimen Types must be assigned to all samples To assign specimen types 1 Open the project 2 For each row select the appropriate setting in the Specimen Category column a Use the default setting of no export for allelic ladders and controls Note Exporting sample types designated as allelic ladders or controls generates an error message b Select the appropriate specimen type for each sample 3 Save the project 9 8 GeneMapper ID Software v3 1 User Guide CODIS Table Export To export the CODIS table 1 Select File gt Export Table for CODIS F Export CODIS Data for lt casework_1 gt __ GeneMapper J Automation Config _ Data Database does _ GM3 1 Database Defaults installer __ JRE _ Size _ Table F File name leasework_1 Export hty Computer Eiai eons formats CMF 1 0 CMF 3 0 dat ox Cancel 2 Complete the Export CODIS dialog box a Navigate to the location you wish to e
49. BENG pes Sample Name Sample Type Specimen Cater Analysis Method Pan aireat 4 C0 Eontr Sample no export None Nan Mone 2 cOladder sample moexport None No None E e Sampe Positive Control moexport None No Mone z E Sample Alale aCe no export Mone Mone merous lLsx _ O COS Negative Control GeneMapper ID Software v3 1 User Guide Analyzing Samples Files Note It is recommended that you choose the appropriate sample type To make it easier use the data collection sample sheet to make these entries refer to Appendix G There is a PQV within the software known as Control Concordance CC If you choose the appropriate sample type for your negative and positive controls the software will automatically check to make sure your negative control does not give any labeled peaks above the peak amplitude threshold and your positive control genotype profile matches the expected profile for 9947A Control DNA If the software finds that your controls match you will get a green flag for the CC You will get a yellow flag for CC if the controls do not match This 1s a quick and easy way to check or double check your controls Note If you use a positive control other than 9947A you can change the control alleles in the marker table within the panel manager to automatically scan for concordance to your control s profile Refer to Chapter 4 Using Panel Manager However you can only have one control
50. Basic Advanced or Classic mode to detect peaks and process data See Basic Mode Peak Detection Algorithm Settings on page B 6 Classic Mode Peak Detection Algorithm Settings on page B 7 and Advanced Mode Peak Detection Algorithm Settings on page B 12 This algorithm uses a dynamic programming approach that is efficient runs in low polynomial time and space and guarantees an optimal solution It first matches a list of peaks from the electropherogram to a list of fragment sizes from the size standard It then statistically derives quality values determined by examining the similarity between the theoretical and actual distance between the fragments Size Matching Algorithm Example An example of how the size matching calling algorithm works 1s shown below using a contaminated GeneScan 120 size standard data Peaks found standard and contamination are indicated by blue lower bars along the x axis The size standard fragments as determined by the algorithm and their corresponding lengths in base pairs are designated by the upper green bars Note that there are more peaks than size standard locations because the standard was purposely contaminated to test the algorithm The algorithm correctly GeneMapper ID Software v3 1 User Guide GeneMapper ID Software Genotyping Algorithms identifies all of the size standard peaks and removes the contamination peaks denoted by the black triangles from considerati
51. Concordance Checks saidas apad oe Aa hae ad emt aed 7 32 Using Plot Windows Samples and Genotypes About the Plot Window 0 ccc eee ee ee ee ees 8 2 Piot Window TOOIDANS lt 0 x obese ele ee i A ae wv eee eae 8 4 Plot Window Menus 00 0c eee ee eee ee ee es 8 5 Exporting Table Data INTFOQUCTION 44 caus wa oe ASO Eee eRe BA 9 2 Exporting Samples and Genotypes Table 000005 9 2 ADOULCODIS asawe et hia eee ON ae a eee eae Re Pee 9 5 CODIS Export Manager 0 0 0 ees 9 6 CODIS Table EXPONE 6 viii saci oom ee widely Se Aled We aan Roe ee ae es 9 8 Printing Data PINTO Dala ede de tay as Wie da Rg hte ai Oat oe ae ee ara 10 2 Page Set Up Options 0 00 eee 10 4 Printing Sizing Plots Similar to Genescan Using the Predefined AmpF STR Sizing Plot Setting 0 0 cee eee 10 7 Printing Genotype Plots Similar to Genotyper Using the Predefined AmpFLSTR Genotyping Setting 000 eee 10 8 Process Quality Values Overview of Process Component Based Quality Values PQV A 2 Using Process Component Based Quality Values A 3 Appendix B Software Genotyping Algorithms GeneMapper ID Software Genotyping Algorithms B 2 Basic Mode Peak Detection Algorithm Settings B 6 Classic Mode Peak Detection Algorithm Settings B 7 Advanced Mode Peak Detection Algorithm Settings B 12 Peak Det
52. D75820 yellow 251 00 298 50 10 11 4 0 090 none 6 7 8 9 10 11 12 13 14 15 Panel cofiler_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 0165539 blue 229 00 279 00 11 12 4 0 130 none Yip Se tp ye 13 14 15 AMEL green 106 00 114 00 x 9 0 000 none XY THOL green 159 00 205 00 8 939 3 4 0 060 none EET e tesa TPOX green 212 00 254 00 8 4 0 060 none iy 7 8 9 10 11 12 137 CSFLPO green 275 00 323 00 10 12 4 0 090 none 6 7 8 9 10 11 12 13 14 15 D75820 yellow 251 00 298 50 10 11 4 0 090 none Diy Fp Se Ap bos desde 5 Panel SGM_Plus_v1 D351358 blue 98 00 148 00 15 16 4 0 110 none 12 13 14 15 16 17 18 19 E 4 Lay Figure 4 2 Example of a panel definition from a spreadsheet saved as a txt document Note There are no extra rows between markers and panels GeneMapper ID Software v3 1 User Guide 4 25 Chapter 4 Using Panel Manager Column Descriptions 1 Marker name locus name 2 Dye color 3 Minimum ASR allele size range actually marker size range 4 Maximum ASR allele size range actually marker size range 5 Control allele names as comma separated bin names 1 2 If none there must be a dash 6 Type of repeat 2 dinucleotide 3 trinucleotide 4 tetranucleotide 5 non repeat 7 Marker specific stutter ratio if none use zero 0 0 8 Comments if none write none 4 26 GeneMapper ID Software v3 1 User Guide Form
53. End value to an increasingly negative number Note The size of a detected peak is the calculated apex between the start and end points of a peak and will not change based on your settings B 26 GeneMapper ID Software v3 1 User Guide Slope Threshold Examples Slope Threshold Examples Initial Electrophero gram Electrophero gram After Adjustments The initial analysis with a value of 0 for both the Slope Threshold for Peak Start and the Slope Threshold for Peak End value produced an asymmetrical peak with a noticeable tail on the right side 2120 2130 2140 2150 2160 2170 2180 2190 2200 2210 2220 2230 2240 2250 2260 2270 2280 2290 2300 2310 2320 1800 _ 1600 Figure B 8 Electropherogram showing an asymmetrical peak After reanalyzing with a value of 35 0 for the Slope Threshold for Peak End the end point that defines the peak moves closer to its apex thereby removing the tailing feature Note that the only change to tabular data was the area peak size and height are unchanged GeneMapper ID Software v3 1 User Guide B 27 Appendix B Software Genotyping Algorithms Ss 2120 2130 2140 2150 2160 2170 2180 2190 2200 2210 2220 2230 2240 2250 2260 2270 2280 2290 2300 2310 2320 Figure B 9 Electropherogram showing the effect of changing the slope threshold for peak end B 28 GeneMapper ID Software v3 1 User Guide Size Calling Methods for Classic and Advanced Modes Size Calling Methods fo
54. Figure B 3 Electropherogram showing two resolved alleles detected as a single peak Note For information on the tick marks displayed in the electropherogram see Examining Peak Definitions on page B 18 B 20 GeneMapper ID Software v3 1 User Guide Optimizing Peak Detection Sensitivity Example 1 Effects of The figure below shows that both alleles are detected after Decreasing the _ re analyzing with the polynomial degree set to 3 while decreasing the Window Size window size value to 15 from 19 data points Value 4140 4150 4160 4170 4180 4190 4200 4210 4220 4230 4240 4250 4260 4270 4280 4290 4300 4310 4320 4930 4340 4350 4360 4370 4980 4390 4400 4410 4420 B ee ee EE EE a a 15 90 185 87 i 15 92 _ 186 78 Figure B 4 Electropherogram showing the alleles detected as two peaks after decreasing the window size value GeneMapper ID Software v3 1 User Guide B 21 Appendix B Software Genotyping Algorithms Optimizing Peak Detection Sensitivity Example 2 Initial The figure below shows an analysis performed using a polynomial Electrophero degree of 3 and a peak window size of 19 data points gram 1820 1830 1840 1850 1860 1870 1880 1890 1900 1910 1920 1930 1940 1950 1960 1970 1980 1990 2000 2010 2020 2030 2040 1800 _ 1600 Figure B 5 Electropherogram showing four resolved peaks detected as two peaks B 22 GeneMapper ID Software v3 1 User Guide Effects of Reducing the Window Size Value and
55. Filter b Ctrl g Overlay All e Combine Dyes All dye colors from a sample are combined into a single source electropherogram ox Samples Plot Fie Edit View Tools Alleles Help Plot Setting HO Genotyping Paresh z ME E a ed AE Fi E ES EE EARN eee X 205 37 Y 1041 GeneMapper ID Software v3 1 User Guide Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued ltem Description Plots e Separate Dyes Each dye color from a sample is continued separated into an individual electropherogram G Samples Plot File Edit view Tools Alleles Help Plot Setting JHID Genotyping x Panes fa z EG Seal E aall AMIE l Fi E ES 0 i D7 820 THOI ID13S317 ID16 539 240 220 23 2 260 D351358 1338 4100 41140 120 130 140 150 4160 170 180 190 200 0 240 250 270 280 290 300 310 320 330 340 350 360 1 tt rete tet 4 tt tt tt ID19 433 FWA TPOX D18551 4100 4140 120 130 140 150 460 170 180 190 200 2140 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 tp tp ttt tt tt 4 A 4 4 4 tthe tt Sample f 4 X 232 49 Y 230 Bin 36 Marker D21511 e Overlay All Multiple samples are combined into a single electropherogram File Edit View Tools Alleles Help Plot Setting Ho Genotyping x Panes
56. GeneMapper ID v3 1 Version GMv3 0 Kit type Microsatellite Chemistry Kit AmpFLSTR_Panels_v1 Panel Blue_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 VWA blue 151 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none 18 19 20 21 22523 24 25 26 26 2 27 28 29 30 Panel Green_I_v1 AMEL green 106 00 114 00 x 9 0 060 none x yT THOL green 159 00 205 00 8 9 3 4 0 060 none 5 6 7 8 9 9 3 10 TPOX green 212 00 254 00 8 4 0 060 none 6 7 8 9 10 11 12 13 CSF1PO green 275 00 323 00 10 12 4 0 090 none 6 Z 8 9 10 11 12 13 14 15 Panel Profi ler_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 VWA blue 151 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none 18 19 20 21 22 23 24 25 26 26 2 27 28 29 30 AMEL green 106 00 114 00 x 9 0 000 none 5 THOL green 159 00 205 00 8 9 3 4 0 060 none 5 6 7 8 9 9 3 10 TPOX green 212 00 254 00 8 4 0 060 none 6 7 8 9 10 11 12 13 csF1PO green 275 00 323 00 10 12 4 0 090 none 6 7 8 9 10 11 12 13 14 15 D55818 yellow 128 00 180 00 LL 4 0 100 none 7 8 9 10 11 12 13 14 15 16 D135317 yellow 192 00 242 00 11 4 0 100 none 89 10 1L 1213 14 157 D75820 yellow 251 00 298 50 10 11 4 0 090 none By F89 10 tl oe iss l4 25 Panel Profi ler_Plus_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 VWA blue 15
57. Manager Applied Biosystems provides three default tables for each application e AmpF STR Default e Microsatellite Default e SNP Default You need to import these default tables into the database through the Table Settings tab GeneMapper ID Software v3 1 User Guide Section 5 3 Table Settings Tab To import a default table 1 Press Import A dialog box opens 2 Double click the Tables Settings folder to display all of the default tables 3 Select the default table you wish to import then click Import The default table appears in the Table Settings tab Note These profiles can be used as models for customizing new profiles Table Settings Editor Introduction When you click New or Open on the Table Settings tab the Table Settings Editor window opens Use the Table Settings Editor to show or hide information specific to your sample add filtering capabilities and give content information for each column in the project window Note You must have a table highlighted to activate the Open button F Table Setting Editor General Samples Genotypes Table Settings Description OK Cancel Figure 5 17 Table Settings Editor The Table Settings Editor window contains the following tabs e General e Samples e Genotypes GeneMapper ID Software v3 1 User Guide 5 29 Chapter 5 Using GeneMapper Manager The General Tab The General tab provides the capability to give a name and descr
58. Peak Detection and Genotyping Test Parameters The test parameters for the peak detection and genotyping category are described below 1 Peak detection evaluation comparing the analysis results from GeneScan software Macintosh OS to GeneMapper ID software using the Classic algorithm and GeneScan software Windows NT OS to GeneMapper JD software using the Advanced algorithm 2 Genotyping evaluation comparing the analysis results from GeneScan software Macintosh OS to GeneMapper ID software using the Classic algorithm and GeneScan software Windows NT OS to GeneMapper JD software using the Advanced algorithm 3 Editing functionality test 4 Locus concordance between duplicate amplifications of the same sample and between AmpF STR kits 5 Matrix and multi component test using Classic and Advanced algorithms 6 Size standard and size match editor using Classic and Advanced algorithms Peak Detection The first parameter evaluated is the comparison of peak height Peak Height differences between GeneScan Software v3 1 2 Macintosh OS to Evaluation GeneMapper JD Classic peak detection algorithm and GeneScan Software v3 7 1 Windows NT OS to GeneMapper JD using the Advanced peak detection algorithm using a subset of the samples described in Table I 1 on page I 4 GeneMapper ID Software v3 1 User Guide l 7 Appendix GeneMapper ID Software Verification Table I 4 displays the results for the combined peak height
59. STR SGM Plus PCR Amplification Kit AmpF STR Profiler Plus ID PCR Amplification Kit AmpF STR Profiler Plus PCR Amplification Kit AmpF STR COfiler PCR Amplification Kit AmpF STR SEfiler PCR Amplification Kit AmpF STR Profiler PCR Amplification Kit AmpF STR Blue PCR Amplification Kit AmpF STR Green I PCR Amplification Kit GeneMapper ID Software v3 1 User Guide 1 3 Chapter 1 About GeneMapper ID Software Unique GeneMapper JD software v3 1 includes the following unique Features of features GeneMapper D Software v3 1 Unique Feature Description CODIS Export The software can export results in a CODIS recognized format cmf v1 0 and 3 0 Process Component Based The PQV system automatically assigns values to the quality of Quality Values PQV the data in respect to sizing and allele calling Poor quality See Appendix A for more samples are those below the user defined thresholds information GeneMapper D Database The GeneMapper D database stores the following data Note The database does not e Predefined and custom designed size standard definitions store individual sample files e Panel marker loci and allele bin definitions e Analysis methods e Table profiles for generating tabular reports e Saved projects with sizing and genotyping results e Matrix files 810 and 377 instruments only e Plot settings Automated concordance checks e GeneMapper ID software compare
60. Sample GeneMapper ID Software v3 1 User Guide C 21 Appendix C Project Window Software Interface Note The Panel and Sample Type properties accounts for close spellings and abbreviations as shown in the following table Case Result A sample file field contains The sample type for the sample is set either Control Ctrl Cri Cntrl to Control or a similar variation Analysis Tab The Analysis tab contains preferences associated with the analysis of data Startup Add Samples Analysis Users Automatic Analysis E Automatically bring low quality samples to the top Quality Metrics Display ie Symbols Numbers If only one labelled allele in a genotype then duplicate the label E Duplicate homozygous alleles OK Cancel Element Description Automatic Analysis When samples are analyzed the software group box sorts the table automatically to bring samples with errors to the top of the table Same functionality as the Bring Errors To Top icon in the Project window Automatically brings errors to the top of the table check box C 22 GeneMapper ID Software v3 1 User Guide Project Window Menus Element Description Quality Metrics Display group box Symbols button Numbers button If only one allele is labeled in a genotype then duplicate the label group box Duplicate homozygous alleles checkbox Display quality
61. Sizing Quality and Precision 5 Click Apply then click OK to close the Size Match Editor and return to the Project window Notice that after overriding the SQ values e The SQ flags are changed to green squares Pass e The Sizing Quality Override SQO column is checked on QO SFNF 5Q XXX XX HE weaeleesle EIEII CILI 6 Ifone ofthe following occurs e Size standard peak assignments are incorrect for one or more of the samples in a subsequent analysis e Fewer than three peaks are matched and a red flag is displayed Define a new size standard for the affected samples and then reanalyze the sample Advanced Basic Peak Detection Algorithm To examine the size standard 1 Check the SQ for all samples Green squares indicate that the samples passed the sizing criteria 2 Ifthe SQ is flagged yellow or red highlight the row for that particular sample and select Analysis gt Size Match Editor 3 Ifthe size standard peak assignments are incorrect for one or more of the samples you can adjust the peak assignments within the size match editor Refer to the next section for more information about the size match editor GeneMapper ID Software v3 1 User Guide 7 13 Chapter 7 Analyzing and Interpreting Data Size Match Editor Introduction The Size Match Editor window enables you to examine size standard electropherograms edit the identification of size standard peaks and view
62. There are six columns for each allele to indicate name size height area mutation and comments SHP Sharp Peak e A flag Pass or Check Ad is automatically displayed when there is a sharp peak present in the marker signal e Analysis does not stop if problems are detected with these properties but you should examine results flagged with Check values e The PQV system gives a label of SHP to indicate a cluster of peaks with a large narrow peak in the middle whose width is 50 less than the neighboring peak e This flag is used for Microsatellite dinucleotide only analysis e Not editable AN Allele Number e A flag Pass Ii or Check A is automatically displayed when the number of alleles exceeds the maximum legal number for the organism or no alleles are found That number is specified in the analysis method e Triggering this flag reduces the final PQV Genotype value GQ parameter to zero O multiplier e Not editable OBA One Basepair Allele e A flag Pass Ii or Check A is automatically displayed when there is a one base pair allele a microvariant peak present in the marker signal e Two allele peaks that are one base pair apart are flagged This may indicate the presence of a microvariant and or an invalid allele call e This flag is used only for Microsatellite dinucleotide markers only e Not editable GeneMapper ID Software v3 1 User Guide A 7 Appendix A Proc
63. Y and Roby R K 2003 Concordance Study of STR Results Using Multiple ABI PRISM Genetic Analysis Instruments AmpFtSTR Kits and ABI PRISM Analysis Software Proceedings of the American Academy of Forensic Sciences Denver Colorado Publication Printers Corp pp 24 25 Leibelt C S Boland C E Brown C L Hatch T L Daoudi Y Lou Y and Roby R K 2003 Verification of GeneMapper JD Software for STR Analysis Proceedings of the American Academy of Forensic Sciences Denver Colorado Publication Printers Corp pp 50 51 GeneMapper ID Software v3 1 User Guide software Warranty This appendix describes the software warranty provided by Applied Biosystems for the GeneMapper ID Software Version 3 1 This appendix covers Computer On MOUraliON ess cereri omen E nae el oa cae wees J 2 Limited Product Warranty 1 0 0 0 00 ccc eens J 2 GeneMapper ID Software v3 1 User Guide J 1 Appendix J Software Warranty Computer Configuration Configuration Applied Biosystems supplies or recommends certain configurations of computer hardware software and peripherals for use with its instrumentation Applied Biosystems reserves the right to decline support for or impose extra charges for supporting nonstandard computer configurations or components that have not been supplied or recommended by Applied Biosystems Applied Biosystems also reserves the right to require that computer hardw
64. a spreadsheet application The Sizing Quality SQ and Genotype Quality GQ display as a numeric value from 0 to 1 The lowest quality is 0 and 1 is the highest value All other PQVS have one of the following values Value in Spreadsheet Definition TRUE Yes there is an X in this cell FALSE No there is not an X in this cell 0 Green pass 1 Yellow check 1 Grey Edited 2 N A Not Applicable GeneMapper ID Software v3 1 User Guide Exporting Samples and Genotypes Table as Marker Dye Allele 1 Allele AE CAE CrADO AE BIN PHR LPH SPU AN BD CC OWL GQ 2 COfiler v1 0391358 B 14 15 FALSE FALSE ooo 0 0 00o 0 0 0 05 3 COfiler v1 D165539 B 11 12 FALSE FALSE co o 0 0 00 of of 0 O86 4 COfler vi AMEL G FALSE FALSE o o 2 0 00o 0 0 0 05 5 COfiler v1 THOI G e 93 FALSE FALSE 0 of of of of of of of of 05 6 COfler vi TPOX G g FALSE FALSE ol of of of of of of of o5 F COfiler v1 CSFIPO G 10 12 FALSE FALSE o of of of of of of of of 05 Ce COfiler v1 D75620 Y 10 11 FALSE FALSE co o 0 0 00 of of 0 O85 9 COfiler v1 0391358 B 12 13 TRUE FALSE ol 2 2 of of of ol 2 olovs 410 COfiler v1 D165539 B 5 z TRUE FALSE ol 2 2 of of of of 2 olove 41 COfiler vy AMEL G FALSE FALSE ol 2 2 of of of ol 2 oog2 12 COfiler v1 THOI G 5 z TRUE FALSE ol 2 2 of of of of 2 of og 13 COfiler v1 TPOX G B 7 TRUE FALSE ol 2 of of of of 2 o o s2 14 COfler v1 C
65. adding new samples automatically Set Analysis Method to JHiD_Advanced for all samples i Read trom the Sample Set Size Standard to 2 ce_F_Hip_essoo r for all samples i Read trom the Sample Set 310377 Matrix to s for all samples ie Read from the Sample Set Panel to Select a Panel for all samples Read from Data collection Comment Panel field Set Sample Type to C Bample r for all samples Read from Data collection Info field OK Cancel Element Description Set Analysis Method to When you add samples the Analysis drop down menu and Method property is set to the value in the text box which contains the list of analysis Read from the Sample methods from the GeneMapper D option button database Set Size Standard to When you add samples set the size drop down menu and standard property to the value in the text box which contains the list of size standards from the GeneMapper D database Read from the Sample option button Set 310 377 Matrix to Set the matrix file to the dye you are using or drop down menu read from the sample file Read from the Sample option button C 20 GeneMapper ID Software v3 1 User Guide Project Window Menus Element Description Set Panel to e Select a Panel for all samples option button and window e Read from the Data collection Comment Panel field option button Wh
66. af wh E es Sample Name Pind sgo sq al Zoomed In View i zj KT D gt x 178 51 Y 974 Note Double clicking with the left mouse button on an axis automatically resets that axis to full view Samples Plot k o x Fie Edit View Tools Alleles Help Piat Setting HD Genotyping AA M resi A E S a a EA E fa a E Sample Name Pand sgo SQ a Full View X and Y Axes o h i z OEA gt x 258 15 Y 1078 8 8 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued ltem Description X Axis Selects the horizontal scale units for the X axis to be Scale displayed in either base pairs default or data points F X Axis Zooming E x ates ns Zooms in on a specific region on the X axis of the electropherogram as follows e Click and drag with the left mouse button the desired region on the X axis In the Samples Plot this action affects all electropherograms In the Genotypes Plot this action affects only the selected electropherogram or e Right click the X axis to open the Zoom To dialog box which enables specific zoom coordinates and tick spacing units to be set Double click the X Axis Sample Plot and all the electropherograms return to Full View Double click the X Axis Genotype Plot and only the selected electropherograms return to Full View A Basepairs Cirl y AXIS Scale t Da
67. and then define the peaks using the Size Standard Editor When editing a predefined size standard only the Size Standard Editor dialog box will be shown To specify size standard parameters l Select Tools gt GeneMapper Manager Then click the Size Standards tab Click the New button to open the Select Dye and Analysis Method dialog box Select Dye and Analysis Method Classic Dye Fed T Analysts Method Advanced HID m Select Sample OK Cancel Perform the applicable procedure below to use the Size Standard Editor Note The mode you select must be the same as the peak detection algorithm you use to analyze the samples in Analysis Method GeneMapper ID Software v3 1 User Guide 5 67 Chapter 5 Using GeneMapper Manager Using the Size To use the size standard editor in classic mode Standard Editor i 1 Complete the Select Dye and Analysis Method window in Classic Mode L Select Dye and Analysis Method Classic Dye Red Analysis Method HD classic T Select Sample CO_Contral fsa OK Cancel a Select Classic b Select a dye color from the drop down list c Select the Classic Analysis Method from the drop down list d Select the row for the particular sample file that contains the dye standard you want to use as the template e Click OK The Size Standard Editor opens 5 68 GeneMapper ID Software v3 1 User Guide Section 5 6 Size Standar
68. bin set was deleted 8 24 GeneMapper ID Software v3 1 User Guide Plot Window Menus Alleles menu items and descriptions continued Item Description Add Allele Ctrl L Allows you to add a new allele call to an unlabeled peak as follows 1 Select Peak Selection mode 2 Select an unlabeled peak using the left mouse button 3 Select the Add Allele menu item or right click mouse button to open the Add Allele dialog box If Allele Edit Comment is selected the Add Allele Comment dialog box opens allowing you to enter an allele comment for the selected peak Labels must be turned on A bin name is assigned to the label if the selected peak falls within a bin defined for the marker Otherwise the allele is designated with a question mark until you rename the allele If the Show type of edit checkbox is selected in the plot profile the word Added displays inside the box Note For quick access to Add Allele Call double click an unlabeled peak Delete Allele Delete Allows you to delete an allele call from a labeled peak as follows 1 Select Peak Selection mode 2 Select a labeled peak using the left mouse button 3 Select the Delete Allele menu item or right click mouse button to open the allele edit options and click Delete If Show Allele Edit Comment is selected the Delete Allele Comment dialog box opens allowing you to enter an allele comment for the
69. choices may be made The Select a Panel window contains the list of panels from the GeneMapper D database When you add samples the panel for all samples is set to the value in the Sample Sheet comment column Note For the 3100 instrument this column is titled Color Comment For the 310 and 377 instruments this column is titled Comment Leave the text alone which selects the existing name or Click the field to present the Select a Panel window to select an existing panel Set Sample Type to e Sample type for all samples option button and drop down menu e Read from Data collection Info field option button When you add samples the Sample Type for all samples is set to the value chosen from the Sample Type drop down menu which contains choices for Sample Control Allelic Ladder and Primer Focus When you add samples the GeneMapper D software does a best guess match comparing the contents of the field specified in the Sample type box with the list of sample types Sample Control Allelic Ladder Negative Control and Primer Focus If a match is found then the Sample Type property for the sample is set to that sample type If a match is not found then the Sample Type property for the sample is set to Sample GeneMapper ID Software v3 1 User Guide Analysis Tab Note The Panel and Sample Type properties accounts for close spellings and abbrevia
70. co Gy BS w LL A ell ES 6 Table Setting it Table Py a Ge Project window toolbar icons and descriptions Icon Description Creates a New Project Tooltip New Project Opens the Open Project dialog box Tooltip Open Project Saves the Project Tooltip Save Project Opens the Add Samples To Project dialog box Tooltip Add Samples To Project Ctrl C Opens the Export dialog box exports the contents of the Samples or Genotypes tables in tab or comma delimited format Ctrl E Tooltip Export Table Reese Opens the Plots Table window Tooltip Display Plots Opens the Size Match Editor window Tooltip Size Match Editor Opens Analysis Method Editor Tooltip Analysis Method Editor Opens the Panel Manager window Tooltip Panel Manager Opens GeneMapper Manager Tooltip GeneMapper Manager Starts the analysis displays progress bar and Stop button on the Status bar during analysis Tooltip Analyze Ctrl R THe 0 E E 3 18 GeneMapper ID Software v3 1 User Guide Navigating in the Project Window Project window toolbar icons and descriptions continued Icon Description Brings samples with lower quality values to the top of the table sorts by overall Genotype quality Tooltip Bring Low Quality Samples to the Top Ctrl B Opens the Table Setting Editor for the currently selected table Tooltip Table Setting Editor Ctrl T Opens t
71. d Click Add the window closes and the samples files display in the new project window GeneMapper ID Software v3 1 User Guide 7 5 Chapter 7 Analyzing and Interpreting Data Applying Analysis Settings Before you can analyze your sample files define the analysis settings in the software The components you must select within the main project window are as follows e Sample Type e Analysis Method e Panel e Size Standard e Matrix 310 and 377 sample files only Note Blank projects can be set up with default analysis settings These defaults can be selected in the Options window under the Tools Menu Refer to Chapter 6 Options Dialog Box for more information In addition these analysis settings can be read from the sample file depending on what information was entered into the sample sheet Refer to Appendix G Sample Sheets and Plate Records To select the appropriate analysis settings 1 Inthe Sample Type column use the drop down list to select one of the following e Sample DNA evidence or database samples e Positive Control e g Control DNA 9947A or Control DNA 007 e Allelic Ladder e Negative Control e Primer Focus for SNaPshot kit only E F GeneMapper ID 3 1 Untitled gmid Is Logged In E ioj x File Edit Analysis Roe S SS E E Froject Casework Progress Status View Tools Help U EE Takie setina fho Tanie MENEE Samples
72. degraded _ki62 fsa D1 Female _32_2ng fsa D2 Male_8_2ng fsa he 7s En dta fom PAE OK Cancel Apply Figure l 14 Size Match Editor using the Advanced algorithm displaying a sizing quality value of 0 9 Algorithm Testing The second category presented is the Algorithm testing Test Parameters The tested parameters are described below l Global cut off value test using the Classic and Advanced algorithms Minus A cut off value using the Classic and Advanced algorithm Off scale flag verification using Classic and Advanced algorithms Off scale display verification using Classic and Advanced algorithms Out of bin allele flag verification using Classic and Advanced algorithms Spectral pull up flag verification using Classic and Advanced algorithms GeneMapper ID Software v3 1 User Guide l 23 Appendix GeneMapper ID Software Verification 10 Le 12 13 Broad peak flag verification using Classic and Advanced algorithms Peak height ratio flag verification using Classic and Advanced algorithms Allele Number flag verification using Classic and Advanced algorithms Low peak height flag verification using Classic and Advanced algorithms Control concordance flag verification using Classic and Advanced algorithms Overall genotype quality verification Overall size quality verification Global Cut off The first parameter tested in the Algorithm testing category is the Value Testing global c
73. determine the PQVs for certain peak metrics The PQV flags green if the actual value for that item is above the user defined threshold A PQV flags yellow if the actual value falls below the user defined threshold These PQVs are used in the calculation of the overall Genotype Quality When you click the Peak Quality tab the following page opens Analysis Method Editor HID Signal level Homozygous min peak height 200 0 Heterozygous min peak height hi 00 0 Heterozygote balance Min peak height ratio lar Peak morphology Max peak width basepairs fi 5 Pull up peak Pull up ratio 005 Allele number Max expected alleles k Factory Defaults OK Cancel Figure 5 14 Peak Quality Tab of the Analysis Method Editor GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab The settings include the following e Signal level in fluorescent units Homozygous min peak height default 200 Heterozygous min peak height default 100 Peaks between the minimum peak height and the signal peak height are sized and called but the peak height flag is yellow and the PQV value will be lowered as a result e Heterozygote Balance Min peak height ratio default 0 5 In a heterozygous allele the higher peak height is X and the lower peak height is Y If the ratio Y X is less than the heterozygote balance then the process quality value flag for peak height ratio will be triggered and the GQ
74. drives or network servers by expanding folders in the navigation pane b Use to table below to add folder s or samples To add Do this A folder containing Select the folder sample files Multiple folders Use the Shift key to select containing sample continuous folders or the Ctrl key to files select discontinuous folders Some of the files ina Use the Shift key to select folder continuous samples or the Ctrl key to select discontinuous samples a The data contained in each run folder is analyzed by comparison of the allelic ladder within that particular folder It is required to have at least one allelic ladder per folder 7 4 GeneMapper ID Software v3 1 User Guide Analyzing Samples Files Add Samples to Project Edit view Files Gh Database Samples To Add biy Computer Fg 3 Floppy 42 Fal Compact Diac Dc EE Local Disk C E E Local Disk E BC 0 AppliedBiosystems E h Config 7 Database e ii Pale Data c Click Add to List The folder will then appear in the right pane Note If you select a couple of sample files the folder from which the samples were selected appears in the right pane GALT i Samples to Project Edit view Files GM Databaze Samples To Add Casework AAS a enh el H E Config ao Database EL eee Data CA HD nam 2 sivas Bo PP Sarnpled ed PP Sample Clea Add a Analyze Cancell Options
75. e LL EL E e rate setna Panels Samples Genotypes H E COfiler Sample Name lRur Na Eel Profiler Plus Sample1 Casework Profiler _Flus_w1 Samplet Casework Profiler Plus v1 a CO_Contral Casework COtiler_v1 4 CO Control ae work ie m 2392 e ooo 5 c0 Control Casework cos vi Soe el TN 6 CO Control Casework COfiler_ v1 CSFIPO LE 10 i 2 1254 T Ca Control Casework Eiler v 0165539 B 1 1 i 2 1484 8 CO Control Casework COfiler_ v1 DIBSS39 B 11 12 1484 E CO Contra Casework Etiler v D351 355 Bo 4 h 5 an 3 1 CO_Contral Casework Eiler v D351 355 BE f 4 5 201 3 h 4 CO_Contral Casework Eiler v 075820 y f 1 1 ert 7 5 mee Tai 3 er 12 La festa hn 1121 1121 1573 1573 1954 1954 Progress gaas ts s s C C C C iCr sCs C C C CC S SCSCsC CS iCidCS ee eee ey et GeneMapper ID Software v3 1 User Guide Appendix K Workflow for Storing Sample Files Projects and Second Reads K 4 GeneMapper ID Software v3 1 User Guide Glossary Allele Allele calling Bin Bin set Diploid polyploid Genotype GM ID HID Kit Marker Microsatellite Panel PQV SNaPshot Kit Multiplex Analysis SNP GeneMapper ID Software v3 1 User Guide A variant of a marker Identification of alleles based on bin definitions genotyping GeneMapper JD software analysis A fragment size or basepa
76. expor HID_Ady Identifiler CE_G5_F Databasir ABIS100 demo_31 12002 06 5 ID_Neg_c ID_Neg_ None Negative no expor HID_Advy Identifiler CE_G5_F Databasit 4BIS100 dermo_31 2002 06 1D Sampi Samples None Sample no expon HID_Advy Identifiler CE_G5_F Databasit 4513100 demo_31 12002 06 ID_Sampl Sample4 Sample no expor HID_Adv Identifiler CE_G5_F Databasit ABIS100 demo_31 2002 06 ID_Sampl Sarmple5 Sample no expon HID_Ady Identifiler CE_G5_F Databasit 4513100 dermo_31 12002 06 rrr amp o io oo iF Analyzing Samples 6 ltem Name Description 1 Drop down These menus are described in Project menus Window Menus on page C 10 2 Toolbar Toolbar icons are described in Project Window File Menu Dialog Boxes on page C 27 3 Samples and The use of these tabs is described in Genotypes Samples View on page C 2 and tabs Genotypes View on page C 7 4 Navigation The use of this pane is described in pane Navigating in the Project Window on page 3 12 5 Samples view These columns are described in Samples columns View Columns on page C 4 6 Progress The progress of analysis and import export indicator of projects is shown in this indicator 3 4 GeneMapper ID Software v3 1 User Guide Parts of the Once you have loaded Sample files into a Project window it looks Project Window like the example below For more information see Appendix C Project Window Software Interface
77. for Search criteria Criteria accumulate as they are entered in different rows and to different groups of criteria Multiple items in the search criteria list use the natural language and for searching For example the natural language of the Criteria field is Display samples whose Panel property contains XXX and display samples whose Collection Instrument property contains YYY GeneMapper ID Software v3 1 User Guide C 43 Appendix C Project Window Software Interface C 44 Element Description Search Criteria Cancel button Enabled when a search is in progress search stops when clicked Note You may not stop the search during data transfer from the GeneMapper D database Search button Searches the database using the criteria list and displays the search results in the Search Results field Search Results field Displays the results of the database search sample files Add To List Adds the selected items from the Results field to the button Samples to Add field Samples To Contains the folders and files that the system will add Add field This field uses the tree viewer interface Options button Opens the Options window Clear button Clears the selected files from the Samples To Add field Add button Adds the Samples to Add list into the Project window Cancel button Closes the Add Samples to Project dialog box
78. genotype quality value will be lower e Peak Morphology Max peak width basepairs default 1 5 e Pull up peak Pull up ratio default 0 05 Considers the ratio of any color peak directly under a major peak Flags spectral pull up quality value as in the genotypes table e Allele Number Max expected allele number default 2 You may want to leave the default setting at 2 so that you are flagged when there is a possible mixture especially in casework If analyzing known mixtures or polyploids enter the maximum number of alleles you expect In order to display all alleles you must enter the correct number of alleles into the table profile e SNP for SNP genotyping application only Double peak default 0 5 See PQV Flags on the Genotypes Page in Appendix A GeneMapper ID Software v3 1 User Guide 5 23 Chapter 5 Using GeneMapper Manager Quality Flags Tab 5 24 PQV Thresholds Quality weights are between 0 and 1 Quality Flag Settings Spectral Pull up fos contral Concordance Broad Peak Lowe Peak Height Out of Bin Allele l Off scale Overlap Feak Height Ratio Paw Thresholds Sizing Quality From p75 to 1 0 From 00ta 0 25 Genotype Quality From jars to 1 0 FromO 0te 0 25 Factory Defaults Figure 5 15 Quality Flags Tab of the Analysis Method Editor The ranges you specify in this section determine the flag assigned to the sizing quality and genotyping quality o
79. identifier v1 FGA R ja 26 o EE E 45 Samples Databasing Identifiler_v1 FGA R 24 26 amp a 46 Samples Databasing Identifiler_v1 THO1 G I 6 4 2l Analysis Completed GeneMapper ID Software v3 1 User Guide Genotype View Columns The table in the Genotypes view can display some of the columns in the Samples view see table on page C 4 plus these unique columns Genotypes view columns and descriptions Column Description Marker Not editable name of the marker associated with the record created in the Panel Manager C 7 Appendix C Project Window Software Interface Genotypes view columns and descriptions continued Column Description Dye From Panel Manager not editable indicates which dye is associated with the marker B blue G green Y yellow R red or O orange Note Not used for SNP Genotyping SNaPshot kit analysis Allele 1 2 etc Not directly editable in the table cell allele calls are editable only in the plot windows The call for Allele 1 Allele calls and names are defined in the Panel Manager as bin names The Allele call text box displays a list of valid calls plus one of the following e OL Off ladder for Human Identification HID applications e for unknown for microsatellite and SNP genotyping SNaPshot kit applications e blank for no allele calls Note
80. if Y EG eb E al Wam E mm EK nF T E es Izle KI X 218 22 Y 936 The Overlay All option can be used to verify size standards as follows 1 Select any number of Samples in the Project window 2 Click iiti Display Plots 3 Turn off all dye color icons except the dye color used for the size standard 4 Click a Overlay All to display size standard electropherograms allowing you to check for outliers 8 18 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued Item Description Tables Controls which table if any is viewed at the bottom of the Sample Plot window Ho Table Ctri amp Table Filter Sizing Table Ctri A w Header Genotypes Table Ctiri iy e No Table Turns off Table e Sizing Table Shows sizing table which displays information for all detected peaks in the selected samples Use the Plot Settings Editor to configure the columns shown in the table Use the Table Filter menu item command to control which rows of data are shown e Genotypes Table The genotypes table as displayed in the Project window Genotypes tab is shown for the selected samples Use the Table Setting Editor to configure the columns shown in the table Table Filter Controls which data rows are displayed in the Sizing or Genotypes Table viewed at the bottom of the Sample Plot window
81. if the sample file name is too big ellipses are introduced in the middle of the name then the length of the name is decreased from the middle to a certain length This way the data can still be printed on one page When this option is selected the whole name is printed and there is no special handling to introduce ellipses and truncate data GeneMapper ID Software v3 1 User Guide 10 5 Chapter 10 Printing Data Plot Tab 10 6 Options Start New Page When this option is selected if the table 1s to be printed after plots the table prints on a new page If not selected then table prints immediately after plots are printed starting on same page as plots if space left and may not fit on that page Honor plots per pane When this option is selected the number of plots selected to be displayed per pane 1 8 prints per page Small When this option is selected the number of plots per pane selected in user interface is ignored and a preset size of the plot small prints Medium When this option is selected the number of plots per pane selected in user interface is ignored and a preset size of the plot medium prints Large When this option is selected the number of plots per pane selected in user interface is ignored and a preset size of the plot large prints GeneMapper ID Software v3 1 User Guide Printing Sizing Plots Similar to Genescan Using the Predefined AmpF4 amp TR Sizing Plot Settin
82. ix How to Obtain More Information 000 eee eee ee ee eee X How to Obtain Support naana na aaaea xi About GeneMapper D Software Overview of GeneMapper ID Software 2000s 1 2 Steps Required to Use GeneMapper ID Software 1 5 Organization of the User Guide anana nanana 1 6 Getting Started with GeneMapper ID Software Hardware Requirements nnna aana aa ee eee 2 2 Installing the GeneMapper ID Software 00 e eee 2 3 Registering GeneMapper ID Software 0 0 eee ees 2 6 Logging Onto and Logging Out of the GeneMapper D Software 2 7 Using the Project Window Main GeneMapper ID Project Window 0 00 eee eee 3 2 Overview of the Project Window 0 00 cee eee eee 3 3 Navigating in the Project Window 0000 eee eee 3 12 Using Panel Manager Panel Manager Overview 0000 cece eee ee eee 4 2 Panel Manager Window Commands 000eeeeeeeeee 4 7 Formats of Panel and Bin Text Files 2 00002 e ae 4 22 GeneMapper ID Software v3 1 User Guide iii Chapter 5 Chapter 6 Chapter 7 Panel Table View 2 anaana ee ees 4 34 Marker Table View si hsiedech cs uh etd at ue sede a itt tee cu sade fe detects Sain wr hanes 4 36 Bins View Microsatellites Only 00 000 cee eee ees 4 39 Using GeneMapper Manager IWATFOGUICUI ON lt 4 5 ashe ie eee ee hee sa ne St neers oie oa aay ts 5 2 sectio
83. lovy_input Profiler_plus_NT Profiler_Plus_v1 FGA B 24 28 Ba E m A Z E w E A 4 gt 60 low input Profiler plus NT Profiler Plus w1 VAVA B 16 17 l Brera el Oe i Analysis Completed Bee aa top Figure l 21 Genotype table displaying PQVs PQVs are calculated using an equation that takes the user defined PQVs to generate the genotype quality score PQVs are weighted from 0 1 O weight equals no effect on the final value and weight equals full effect The individual PQVs were evaluated using the default threshold values GeneMapper ID software If a sample meets the individual PQV threshold defined in the software it is treated as a zero in the calculation See examples below Overall Genotype Quality GQ PQV Green 0 75 to 1 0 Yellow 0 26 to 0 7 Red 0 0 to 0 25 The calculation for the Overall GQ PQV by the software is as follows GQ 1 PQVos 1 PQVgm 1 PQVpnr L PQVLpp 1 PQVspu L PQVan 1 PQV3p L PQVce PQVovi GeneMapper ID Software v3 1 User Guide Results of Verification Testing Example A The LPH PQV has a weighted value of 0 3 in the default settings GQ 1 1 0os 1 Opin 1 Opy 1 0 31pu 1 Ospy 1 O4n C Ogp 1 0cc C Yovz l log gm pur 0 3Lpu 1spu lan lep lce love GQ 1 0 3 0 7 An overall GQ PQV of 0 7 displays a yellow flag Example B The PHR and LPH PQVs have weighted values of 0 3 in the default setting GQ
84. marker exhibit a grey triangle to indicate the marker has been modified The genotype quality GQ will show a green square Allele edits made in the plot windows are immediately reflected in the genotypes table When you edit an allele the software stores a log of the changes that were made in the Allele History To edit allele calls Note You must be displaying peaks with labels to edit allele calls l 2 Select the peak by clicking it and right click a Ifthe peak is already labeled you have the option to Delete the allele call Rename Allele or view the History b Ifthe peak is not labeled you have the option to Add Allele Call or view History Choose the appropriate option by clicking on it If you are renaming or adding an allele call choose the allele call or enter a custom name After you make the necessary change you will be prompted to enter an allele edit comment Enter a comment and press OK The changes are automatically be reflected in the genotypes table GeneMapper ID Software v3 1 User Guide 7 25 Chapter 7 Analyzing and Interpreting Data To view the Allele History 1 Select the peak by clicking it and right click 2 Select History to open the Allele History Window You can see all the changes made to the allele There are 3 types of actions that are displayed Created 1s the action in the first row if the peak was labeled by the software automatically The comment al
85. menu items as described throughout this chapter Position your mouse over an icon to view its tooltip description 1 2 3 4 5 6 fiicrscatcun os AIC EE Gee Bir Ma Bae fbi 2 fe de A ee Plot window toolbar icons and descriptions ltem Description 1 Pull down menu that allows selection of a plot profile Note Click the Plot Settings Editor icon to edit the selected profile Tools menu 2 Pull down menu that controls the number of electrooherogram panes shown 3 Dye color toggle icons View menu 4 Header Marker Range and Marker Indicator toggle icons View menu 5 Icons that select the plot display Combine Dyes Separate Dyes Overlay All View menu 6 Icons that select the Sample Plot window to be displayed No Table Sizing Table Genotypes Table View menu T Icons that select either Peak Selection Mode or Binning Mode Alleles menu 8 Icons that select the Label display No Labels Horizontal Labels Vertical Labels View menu 9 Icons toggle additional view features Overlay Bins Show Hide Peak Position Controls To Top Show Hide Allele History View menu 10 Icon switches view to full X axis and Y axis scales Full View from the View menu 8 4 GeneMapper ID Software v3 1 User Guide Plot Window Menus Plot Window Menus File Menu The File menu commands are used to perform the basic commands gli Edit View Tools Alleles H cave Panel P
86. on each window for example File Edit View provide a set of commands specific to the window This section will describe the menu commands and dialog boxes associated with a given window Samples View The following information is provided in this section for the Samples view Topic See Page Appearance of Samples View C 3 Samples View Columns C 4 Samples View Features C 6 C 2 GeneMapper ID Software v3 1 User Guide Overview of the Project Window Appearance of Samples View The Samples view of the Project window displays the samples you want to analyze or have already analyzed This view looks much like the following example when sample data has been imported GeneMapper ID 3 1 Database gmid Is Logged In File Edit Analysis View Tools Help i Fr G z e s e LU l U E mm Table setting amprLstr Table m8 8 Project Samples Genotypes Aetabasing Status Sample Name Sample Type Specimen Cater Analysis Method Panel Size Standard Matrix Run Name lIn ID_Cortral Positive Control no export Identifiler Identifiler_ 1 LIZ_size_standard Databasing A ID_Ladder Allelic Ladder no export HID_Advanced Idertifiler_ 1 LIZ_size_standard Databasing 14 3 ID_Neg_Crirl Negative Contre no export HID_Advanced dentifiler_v1 LIZ_size_standard Databasing 14 4 Samples Sample Convicted Offel HID_Advanced dentifiter_v1 LIZ_size_standard Databasin
87. peak is Delete selected Change Size Opens a dialog box for the Enabled when a Label selected peak allowing the labeled peak is selection of a new size label selected Delete All Removes deletes all size Always enabled Size Labels labels for the selected size Ctrl Delete standard sample 7 16 GeneMapper ID Software v3 1 User Guide View Menu Size Match Editor The View menu is used to change the Size Match Editor window Y Axis Scale show Navigator Ctrl Shittth View menu items and descriptions Item Description Enabling Zoom Submenu used to scale the plot to the following value Full View Ctrl Always enabled Y Axis Scale e Scale to maximum Y default e Scale to user defined Always enabled Show Navigator Ctrl Shift N Switches the navigator pane Removes the pane with the first use and restores it on the second use Always enabled Tools Menu Auto Adjust Sizes Check Sizing Quality Tools menu items and descriptions Item Description Enabling Auto Adjust Auto adjusts size standard Always enabled Sizes peaks Tooltip Auto Adjust Check Sizing Checks the quality of the Always enabled Quality current size matching Tooltip Check Quality GeneMapper ID Software v3 1 User Guide 17 Chapter 7 Analyzing and Interpreting Data 18 Toolbar Using the Size Match Ed
88. sample sheet G 2 Sample Sheet how GeneMapper uses G 2 Samples and Genotypes views applying a menu selection 3 10 purposes of tables 3 5 3 11 Samples view appearance C 3 columns C 4 Samples View column filtering settings 5 36 Save Project Save Project As dialog boxes C 29 second reads workflow for storing K 1 Services and Support obtaining xi setting CODIS export fields 9 6 setting options 6 3 size calling curve black curve 7 19 red curve 7 19 Index 5 size calling method Cubic Spline Interpolation B 31 Global Southern B 34 Local Southern B 32 size calling methods classic and advanced B 29 Cubic Spline B 31 Least Square B 29 Size Match Editor Edit Menu 7 16 icon 7 11 procedure for editing apeak 7 18 View Menu 7 17 when to use 7 14 Size Match Editor window editor commands 7 16 elements 7 15 introduction 7 14 toolbar 7 18 using 7 18 size standard 350 All size standard D 4 400 HD D 5 500 All D 7 creating and editing 5 67 incorrect peak assignments 7 13 specifying size standard parameters 5 67 Size Standard Editor window using in basic or advanced mode 5 71 using in classic mode 5 68 Sizing Quality SQ overriding 7 12 viewing 7 11 7 13 Sizing Quality Override SQO 7 13 slope threshold B 27 peak end parameters B 25 peak start B 25 software setup setting options 6 3 Sort Dialog box C 46 Source Lab ID adding 9 7 selecting 9 9 specimen number CODIS 9 8 specimen type 9 7 SQ and GQ qualit
89. scale display verification the out of bin flag verification the spectral pull up flag verification and the broad peak flag verification using both the Classic and Advanced modes The weighted values for the individual PQVs were evaluated using the default threshold settings in GeneMapper JD Software v 3 1 to verify that the quality score given for each PQV is an accurate assessment of the sample A total of 2772 alleles comprising three 3 off scale samples were evaluated The PQV flags for the off scale samples were presented accurately in the Classic mode However when these same off scale samples were analyzed in the Advanced mode it was observed that the PQV flags for the pull up peaks that were caused by off scale peaks 1 e in the same position in a different color see Figure I 17 are overridden and reflect the PQV flags for the true off scale peak in that position It is recommended that an analyst carefully review all off scale yellow PQV flags since it is known that the flags for the pull up peaks in the same position as the true off scale peak do not accurately reflect the pull up peak in the Advanced mode For example in Figure I 18 the spectral pull up peak is in the same position in a different color of an off scale allele and the off scale spectral pull up and broad peak values do not accurately reflect the PQV flags that are displayed for that position That is the off scale PQV flag is yellow this spectral pull up
90. the base pair distance to be subtracted from the location e g 0 5 4 Bin right offset The right boundary of the bin expressed as the base pair distance to be subtracted from the location e g 0 5 Row Description 5 GeneMapper D version number 6 Kit name Name of chemistry i e AmMpFLSTR_Panels_v1 7 Bin set name The collection of all AmpFZSTR bin sets 8 Panel name An AmpF STR kit 9 Marker name The locus name 4 28 GeneMapper ID Software v3 1 User Guide Formats of Panel and Bin Text Files Panel Definitions The examples below show SNP genotyping panels defined in the for SNP format used by GeneMapper JD software v3 1 in a tab delimited text Genotyping file txt using Notepad or WordPad or an Excel spreadsheet saved as a txt file E SNPdemo txt Notepad a loj x File Edit Format Help wersion GM w 3 1 kit type SNF Chemistry Kit SNPdemo none Fane 6plexdemo none none Column Description 1 Marker name 2 Control allele names as comma separated bin names A G if none there must be a dash 3 Comments if none write none GeneMapper ID Software v3 1 User Guide 4 29 Chapter 4 Using Panel Manager E4 Microsoft Excel Book 3 A O x File Edit View Insert Format Tools Data window Help x bplexdemo none none nore nore none nore nore sheet Sheet y sheda y 4 gt
91. then duplicate the label group box Duplicate homozygous alleles checkbox If this option is checked single alleles will be displayed in the Genotypes table and in the Plot window as two labels to indicate presumed homozygotes GeneMapper ID Software v3 1 User Guide Users Tab Users Tab The Users tab contains preferences associated with managing registered users Lange Passvvorg Note Once created user names cannot be deleted GeneMapper ID Software v3 1 User Guide 6 9 Chapter 6 Options Dialog Box 6 10 Element Description User Name field Displays the list of registered users e User Name column Your name e Created On column Date user was created e Show column Shows your name in User Name drop down menu of the login window if checked Clearing a Show column check box for a user blocks login for that user can be done by anyone running GeneMapper D software Shift click the column label headers to sort the list by that column New User button Opens the New User dialog box Pm 2 Hew User oe Mew username Password Confirm Password To add a user name for GeneMapper D software click New User to open a dialog box and then a Type a user name into the dialog box b Enter a password using only alphabetic characters c Confirm the password d Click OK to assign the user name and close the dialog When you sele
92. user can access a database at a time GeneMapper ID Software v3 1 User Guide 2 9 Chapter 2 Getting Started with GeneMapper ID Software Logging Out Close the GeneMapper ID software in one of three ways e Select File gt Logout e Select File gt Exit or e Click the Close button When you log out of the GeneMapper ID software you have the option of saving or discarding the changes you made since the last time you updated the project Save Project Ed AN Do you want ta save changes Mo Cancel Note If you close GeneMapper JD software with a new or blank Project window or no changes in your Project the Save Project logout dialog box does not open 2 10 GeneMapper ID Software v3 1 User Guide Using the Project Window This chapter provides a general overview of the Project window the central window of the GeneMapper ID Software Version 3 1 interface This chapter covers Main GeneMapper JD Project Window 000 00 ee 3 2 Overview of the Project Window 0000 ee eens 3 3 Navigating in the Project Window 0 0 0 0 eee 3 12 GeneMapper ID Software v3 1 User Guide 3 1 Chapter 3 Using the Project Window Main GeneMapper D Project Window Importance of the The Project window is the primary window in the GeneMapper ZD Project Window software user interface The GeneMapper ID software is organized around the concept of the project to support its purpose of automati
93. values in project tables as symbols green square yellow triangle red octagon or numerical values 0 0 1 0 Note If Symbols default is chosen then all quality values are shown as symbols If Numbers is chosen then only SQ and GQ columns are shown as numbers the other result columns are symbols If this option is checked single alleles will be displayed in the Genotypes table and in the Plot window as two labels to indicate presumed homozygotes GeneMapper ID Software v3 1 User Guide C 23 Appendix C Project Window Software Interface Users Tab The Users tab contains preferences associated with managing registered users Startup Add Samples Analysis Users GeneMapper Users Created on Show Thange Password Note Once created user names cannot be deleted Element Description User Name field Displays the list of registered users e User Name column Your name e Created On column Date user was created e Show column Shows your name in User Name drop down menu of the login window if checked Clearing a Show column check box for a user blocks login for that user can be done by anyone running GeneMapper D software Shift click the column label headers to sort the list by that column C 24 GeneMapper ID Software v3 1 User Guide Project Window Menus Element Description New User button Change Password Opens the N
94. 0 360 a0 0 hs aj b b fu 22 Sangles Identifme v1 C DESB FWA TPOX Disss1 110 4120 130 440 150 460 170 180 190 200 2140 220 230 240 250 260 270 280 290 300 3140 320 330 340 350 360 e D55818 FGA 410 120 4130 440 1450 460 4170 4180 190 200 240 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 BAEN Ki TE X 223 82 Y 121 Bin 34 Marker D21511 e Headers Turned Off File Edit View Tools Alleles Help Plot Setting HID Genotyping zi ranes 11 Ma Efi al SM O ed D851179 D21511 D75820 CSFIPO a 410 120 130 4140 150 4160 4170 180 190 200 2140 220 230 24 250 260 270 280 230 300 310 320 330 340 350 360 D31358 THO D13S317 ID16 539 D2 1338 220 230 240 250 260 410 1420 130 4140 150 4160 170 180 1390 200 210 Si 270 280 290 300 310 320 330 340 350 360 DIss433 FWA TPOX D18551 DIESE M 110 120 130 440 150 4160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 Sa n A S A S A AADA AA St tt ee D55818 FGA 410 120 130 1440 150 4160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 FAKI Ls E 292 98 Y 192 Marker Controls whether or not the colored marker bar is Range displayed at the top of an electropherogram Ctrl R 8 20 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued It
95. 0 sizing precision reproducibility Size Standard GeneMapper ID Software v3 1 User Guide 7 19 Chapter 7 Analyzing and Interpreting Data To check size precision 1 Highlight all the samples in the main project window Edit gt Select All 2 Display the sample plots Analysis gt Display Plots e Ifyou have imported the factory provided plot settings Refer to plot setting sections of Chapter 5 select Overlay LIZ or Overlay ROX depending on the dye color of your size standard from the plot setting drop down box e Ifyou did not import the factory provided plot settings Select only the color of the size standard in the toolbar Select View gt Plots gt Overlay All select View gt Tables gt Sizing 3 Zoom in on the 250 bp peak a Place the cursor to the left of the 250 bp peak along the top x axis until the cursor changes to a magnifying glass b Click and drag to create a box and release when the box includes the 250 bp peak 4 Select the 250 bp peak by clicking it 5 Select View gt Table Filter gt Show Selected Rows 6 Verify that the values within the size column are within bp Note The 250 bp peak should size consistently and overlap In a typical run the 250 bp peaks all fall within a size window of approximately 1 bp Temperature fluctuations in the lab may cause variations gt bp 7 20 GeneMapper ID Software v3 1 User Guide Size Calling Curve Overlay LIZ Use Overla
96. 1 12 TPOX yellow 716 99 260 99 E 4 0 05 none 67881011 4 or 4 F OK Cancel Apply This figure shows the columns in the AmpF STR marker table They are further described in the following table me SS Reference Samples Table 4 3 Marker table column descriptions Column Description Marker Contains the name of the marker Cell accepts Name alphanumeric characters from the keyboard The field is accepted if the marker name is unique in the panel Editable Note Marker names are case insensitive i e D1S83 is the same as d1s83 Dye Color Contains the color names blue green yellow red orange The field is accepted if it is one of these text strings which are not case sensitive Editable 4 36 GeneMapper ID Software v3 1 User Guide Marker Table View Table 4 3 Marker table column descriptions continued Column Description Min Size Contains the starting base pair number for the allele size range Cell accepts numeric characters from the keyboard Editable The range defined by the Minimum Size parameter should include the stutter peaks for the smallest allele Max Size Contains the ending base pair number for the allele size range The cell accepts numeric characters from the keyboard Editable Control Contains a comma separated list of control allele names Alleles in the marker Editable For AmpF STR data analysis the control alleles correspond to Control
97. 1 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none 18 19 20 21 22 23 24 25 26 26 2 27 28 29 30 AMEL green 106 00 114 00 x 9 0 000 none Ky D851179 green 118 00 183 50 13 4 0 120 none 8 9 10 11 12 13 14 15 16 17 18 19 D21511 green 184 50 247 50 30 4 0 130 none 24 2 25 26 27 28 28 2 29 29 2 30 30 2 31 31 2 32 3Zz D18551 green 264 49 350 00 15 19 4 0 160 none 9 10 10 2 11 12 13 13 2 14 14 2 15 16 17 18 19 20 2 D55818 yellow 128 00 180 00 11 4 0 100 none Br ea ree 8 a Ss Bs Uc se eB D135317 yellow 192 00 242 00 11 4 0 100 none 8 9 10 11 12 13 14 15 D75820 yellow 251 00 298 50 10 11 4 0 090 none 6 7 8 9 10 11 12 13 14 15 Panel cofiler_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 0165539 blue 229 00 279 00 11 12 4 0 130 none 5 8 9 10 11 12 13 14 15 AMEL green 106 00 114 00 x 9 0 000 none ae THOL green 159 00 205 00 8 9 3 4 0 060 none 5 6 7 8 9 9 3 10 TPOX green 212 00 254 00 8 4 0 060 none 6 7 8 9 10 11 12 13 CSF1LPO green 275 00 323 00 10 12 4 0 090 none 6 7 8 9 20 11 12 13 14 15 D75820 yellow 251 00 298 50 10 11 4 0 090 none 6 7 8 9 10 11 12 13 14 15 7 Panel SGM_P lus_v1 D351358 blue 98 00 148 00 15 16 4 0 110 none 12 13 14 15 16 17 18 19 Figure 4 1 Example of a Panel Definition in Tab Delimited Text Format Note There are no extra rows between markers and panels GeneMapper ID Software v3 1 User Guide 4 23
98. 1 User Guide Peak Detection Polynomial Degree and Peak Window Size Parameters Effects of In the figure below both polynomial curves have a degree of 3 and Increasing the the window size value was increased from 15 red to 31 black data Window Size _ points Value As the cubic polynomial is stretched to fit the data in the larger window size the polynomial curve becomes smoother Note that the structure of the smaller trailing peak is no longer detected as a distinct peak from the adjacent larger peak to the right 2000 1500 Window size value of 31 black 1000 Window size value of 15 red 500 Figure B 2 Electropherogram showing the same peaks as in Figures B 1 after increasing the window size value while keeping the polynomial degree the same GeneMapper ID Software v3 1 User Guide B 19 Appendix B Software Genotyping Algorithms Optimizing Peak Detection Sensitivity Example 1 Initial The figure below shows two resolved alleles of known fragment Electrophero lengths that differ by one nucleotide detected as a single peak The gram analysis was performed using a polynomial degree of 3 and a peak window size of 19 data points 4140 4150 4160 4170 4180 4190 4200 4210 4220 4230 4240 4250 4260 4270 4280 4290 4300 4810 4820 4830 4340 450 4360 4970 4980 4990 4400 4410 4420 SALN A A 1 Palpa EO A E E E A A E E E E E E E E rede 2750 2500 2250 2000 1750 1500 _ 1250 1000 750 _ 500
99. 3 Appendix J Software Warranty J 4 DATA FAILURE OR INTERRUPTION IN THE OPERATION OF ANY EQUIPMENT OR SOFTWARE DELAY IN REPAIR OR REPLACEMENT OR FOR LOSS OF REVENUE OR PROFITS LOSS OF GOOD WILL LOSS OF BUSINESS OR OTHER FINANCIAL LOSS OR PERSONAL INJURY OR PROPERTY DAMAGE No agent employee or representative of Applied Biosystems has any authority to modify the terms of this Limited Warranty Statement or to bind Applied Biosystems to any affirmation representation or warranty concerning the product that is not contained in this Limited Warranty Statement and any such modification affirmation representation or warranty made by any agent employee or representative of Applied Biosystems will not be binding on Applied Biosystems unless in a writing signed by an executive officer of Applied Biosystems This warranty is limited to the buyer of the product from Applied Biosystems and is not transferable Some countries or jurisdictions limit the scope of or preclude limitations or exclusion of warranties of liability such as liability for gross negligence or wilful misconduct or of remedies or damages as or to the extent set forth above In such countries and jurisdictions the limitation or exclusion of warranties liability remedies or damages set forth above shall apply to the fullest extent permitted by law and shall not apply to the extent prohibited by law GeneMapper ID Software v3 1 User Guide Workflow for St
100. 306 79 Y 1174 Figure l 2 Female single source DNA Sample amplified with the AmpF STR Identifiler PCR Amplification kit electrophoresed on an 310 Genetic Analyzer Macintosh OS then analyzed with GeneMapper ID software using the Classic algorithm GeneMapper ID Software v3 1 User Guide l 11 Appendix GeneMapper ID Software Verification EA Genotyper 3 7 Plots IDENTIFILER_ 1 gta al i File Edit Analysis Category Table Views Macro Window Help 100 120 140 160 180 200 220 240 260 280 300 320 340 360 380 400 420 fA Female_32_1ng fsa 4 Blue Female_32_1ng 7 el Haj 13 m 4 19 2007 2483 2131 1850 1624 1743 2025 Fd Female_32_1ng fsa 4 Yellow Female_32_1ng FA Female_32_ing fsa 4Red Female_32_ing 3000 2000 Ei mj 13 E 23 3409 1929 1659 1682 1346 Figure I 3 Female single source DNA Sample amplified with the AmpF STR Identifiler PCR Amplification Kit electrophoresed on an 310 Genetic Analyzer Windows NT then analyzed with GeneScan v3 7 1 software and Genotyper v3 7 el je ed je 2000 1500 1000 500 g 14 29 21 10 10 12 2198 2215 2022 1417 1462 11 1265 1262 FA Female_32_ing fsa 4 Green Female_32_ing 2000 1000 1500 1000 500 1000 l 12 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Samples Plot la x File Edit View Tools Alleles Help Plot Setting HID Genotyping 7 Panes fa ai EE Ba zs PU aal Pa a
101. 7 3 gmid 377 analyzer HID Mew Open Save Ag Import Export Figure 5 6 Analysis Methods Tab of the GeneMapper Manager Analysis Methods Table 5 4 Description of elements on the Analysis Methods tab Tab Elements ltem Description Name column Analysis method name Last Saved Date Time stamp showing when the analysis column method was last saved Owner column User name of the person who created the analysis method Instrument column The type of instrument used GeneMapper ID Software v3 1 User Guide 5 9 Chapter 5 Using GeneMapper Manager Table 5 4 Description of elements on the Analysis Methods tab Item Description Analysis Type column Identifies the analysis mode of the analysis method e HID Microsatellite SNP Genotyping New button Opens the New Analysis Method dialog box Always enabled This dialog box asks you to select the analysis type before opening the Analysis Method Editor The analysis type you select e Sets the analysis algorithm e Displays the appropriate fields for that type of analysis in the Analysis Method Editor dialog box Open button Opens the editor for a selected analysis method Enabled when a analysis method is selected Save As button Displays the Save As dialog box Enabled when an analysis method is selected Import button xml file type Displays a dialog box for Importing analysis m
102. A typical number might best number for the be 3 for microsatellites daa or 10 for SSCPs l Half width Under denaturing conditions certain fragments in the GeneScan 2500 standard display as doublets or split peaks This standard has labels on both stands of the DNA Under poor denaturing conditions you see split peaks One of the two fragments typically has normal mobility while the other does not The Split Peak Correction feature allows the software to correctly call each of the splits If you select split peak correction you will also need to verify or change the correction limits After you decide on a split peak correction method use the same method for all projects to keep size calling consistent Note Split Peak correction is not used with AmpF STR or SNaPshot applications ltem Description No Peak No correction for doublets Correction GS2500 Peak Makes the following peak size assignments for Correction GeneScan 2500 e The right peak for all fragments 222 233 238 286 and 490 e The left peak for all other splits GeneMapper ID Software v3 1 User Guide Classic Mode Peak Detection Algorithm Settings Item Description Left Most Peak Correction Chooses the left peak for every doublet Right Most Peak Correction Chooses the right peak for every doublet Correction Limit Set a correction limit if correcting for doublets Set this value s
103. Chapter 4 Using Panel Manager Panel Manager Overview 4 2 Introduction Related Definitions Use the Panel Manager utility to manage all of the chemistry kits panels markers loci and bin definitions required for automated allele calling Topic See Page Related Definitions 4 2 Purpose of the Panel Manager Window 4 3 When to Click the Apply Button 4 3 Displaying the Panel Manager Window 4 4 Hierarchy of Panel Data 4 4 Panel Manager Toolbar 4 5 Panel Manager Navigation Pane 4 6 There are two ways to add information to the Panel Manager e Creating kits panels markers loci and bins manually e Importing preformatted text files that contain panels markers loci and bin definitions See Formats of Panel and Bin Text Files on page 4 22 for two examples The words kit panel marker locus bin and bin set are defined in the following table Term Definition Panel Collection of loci A primer set specific to an AmMpFZSTR kit For example Identifiler has 16 loci Kit Collection of panels e g a panel is an AmMpF4STR kit Marker A locus or primer pair For example THO1 or vWA Bin An expected location for a particular allele within a locus Bin Set Collection of expected locations for loci contained within a panel or kit GeneMapper ID Software v3 1 User Guide Panel Manager Overview Purpose of the The Panel Manager window enables y
104. Check range is the interval between Pass and Low Quality if any For example if you use the default threshold settings shown in the figure above any result over 0 75 passes any result at 0 25 or below is of low quality and any result from 0 26 to 0 74 will be marked for you to check See Appendix A Process Quality Values When the default setting for the Quality Metrics Display Analysis tab in Preferences window is Symbols a Pass for the SQ and GQ parameters is indicated by a green square a Check is indicated by a yellow triangle and a Low Quality is indicated by a red octagon Some quality metrics are either Pass or Low Quality for example Background Peak and some are warning flags with either Pass or Check values for example File Not Found Such values are not adjustable by the Threshold parameter GeneMapper ID Software v3 1 User Guide 5 25 Chapter 5 Using GeneMapper Manager 5 26 GeneMapper ID Software v3 1 User Guide Section 5 3 Overview Section 5 3 Table Settings Tab Table Settings Tab Use the Table Settings tab to perform the following tasks e Create new table profiles for viewing and exporting e Hide and show table columns e Filter the table entries rows in Project windows The Table Settings tab allows you to determine what columns are viewed in the Samples and Genotypes tab and set filtering properties The Table Settings tab contains a list of Table Settings in th
105. CheckS s gu dkad eae Dies rice aweictk ees 7 32 GeneMapper ID Software v3 1 User Guide 7 1 Chapter 7 Analyzing and Interpreting Data Introduction The previous chapters described how to customize the software to meet your analysis needs Once the software is configured to analyze a particular set of samples such as 3100 four dye AmpF STR sample files you can use the information in this chapter to analyze your data view the data and interpret PQVs Considerations for HID Analysis When using GeneMapper ID Software version 3 1 to perform Human Identification HID analysis with AmpF STR kits consider the following HID analysis requires the presence of at least one allelic ladder sample per project Multiple ladder samples can be used in an analysis provided individual laboratories conduct the appropriate validation studies For multiple ladder samples suitable processing of allelic bins is carried out whereby an average of all ladders within a run folder using the same panel is used to calculate allelic bin offsets Allelic ladder samples in a single run folder are considered to be from a single run When multiple run folders are imported into a project only ladders within a single run folder are used for calculating allelic bin offsets and subsequent genotyping Allelic ladder samples need to be identified as Allelic Ladder in the Sample Type column in a project Failure to make this setting for ladder
106. D 3 1 casework_1 gmid Is Logged In File Edit Analysis View Tools Help cet cae att LU GA OE Eh Project Samples Genotypes Eil casework Sample Type Specimen Ce j Sample poexpot ampe re export 5 Sample ino export Alec Ladder no export 7 Sample ino export cS O Analysis Completed GeneMapper ID Software v3 1 User Guide Interpreting Sample PQVs Sizing Quality and Precision Interpreting Sample PQVs Sizing Quality and Precision Once analysis is complete check the Sample PQVs process quality values and the Sizing Quality SQ These flags can be found in the samples tab of the main project window For an explanation of the sample PQVs refer to Appendix A Process Quality Values Note The PQV displayed is determined by the Table Setting you have selected Sample PQVS e Ifasample PQV is flagged green the sample file is considered good quality in respect to that sample PQV e Ifa sample PQV non SQ is flagged yellow look at the raw data to determine the problem To look at the raw data from the samples tab in the main project window highlight the sample and select View gt Raw Data If you determine this sample should not be considered for interpretation due to problems within the raw data you can remove this sample file from the project Return to the main project window by clicking the Project node in the left navigation pane The sample will still be highlighted Th
107. D Software v3 1 User Guide Using Process Component Based Quality Values Using Process Component Based Quality Values The PQV System Quality values reported by the GeneMapper JD software are an aid to finding and fixing problems in sample preparation and analysis These values are the end results reported by the PQV system For more information on the PQV system see Unique Features of GeneMapper ID Software v3 1 on page 1 4 The color coded sortable PQV values on the Samples and Genotypes pages allow you to isolate problem samples quickly and examine the electropherograms Two types of PQV values are presented on the Samples and Genotypes pages On the Samples page e The SFNF MNF SNF and OS parameters report results as Pass I or Check e The second type the Sizing Quality SQ parameter reports results as Pass Check or Low Quality B On the Genotypes page e The SHP AN AE OBA SPA Bin PHR LPH SPU BD SP OS CC OVL NB and DP parameters report results as Pass BE or Check e The second type the Genotype Quality GQ parameter reports results as Pass MJ Check or Low Quality F Note On the Genotypes page the ADO and AE parameters are also PQV parameters but they report results as checks gt instead of colored flags A red octagon for the Sample SQ indicates that the sizing standard is of low quality A red octagon for the Sample GQ parameter means the analysis of that sample
108. DNA 9947A or Control DNA 007 Marker Indicates a di tri or tetra nucleotide repeat Repeat Contains a number for the number of repeats 2 3 4 The field also accepts 9 for nonrepeat containing markers The field is valid if it contains 2 3 4 or 9 any other number is invalid Editable Marker Column provided for AmpF STR markers Contains the Specific ratio of stutter peak to main peak Sets the maximum Stutter Ratio value of stutter for a specific marker as opposed to using a global value for all markers Editable Comments Contains comments regarding the marker Cell accepts alphanumeric characters from the keyboard Editable Ladder Column provided for the allelic ladders that come with Alleles the AmpF STR kits Comma separated list of allele names in the allelic ladder Applicable only to Human Identification HID genotyping or applications that use allelic ladders Editable GeneMapper ID Software v3 1 User Guide 4 37 Chapter 4 Using Panel Manager 4 38 Editing Marker Tables To edit information in the Marker Table view highlight the cell you want to edit and enter the new information IMPORTANT If you are going to make changes to one or more of the AmpF STR panels duplicate the panel s first then make changes to the copy If you forget to make a copy you can export and reimport the AmpF STR panels and bins to restore the recommended default values this procedure requires you
109. Edit the bin information in the dialog box that opens For SNP Genotyping data 1 Select a SNP genotyping panel in the Panel Manager navigation pane 2 Select a bin to be edited in the Plot tab 3 Select the Edit Bin menu item or right click the bin and select the Edit Bin option 4 Edit the bin information in the Edit SNP Marker dialog box that opens 4 14 GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands To quickly resize a bin select a bin and adjust the right and left handles to the desired size range To move a bin select a bin and holding down the mouse button move the bin to the desired location Edit Bin Dialog Box x Name 313 Location 12 74 Left offset 0 40 New offset default Right offset io 40 New offset default bao bao Mutant Bin OK Cancel Table 4 2 Edit Bin dialog box elements and descriptions Element Description Name Editable name of the bin can be alphanumeric Location Editable location of the bin center in base pairs Left Offset Editable the left boundary of the bin expressed as the base pair distance to be subtracted from the location default 0 4 Right Offset Editable The right boundary of the bin expressed as the base pair distance to be added to the location default 0 4 GeneMapper ID Software v3 1 User Guide 4 15 Chapter 4 Using Panel Manager Table 4 2 Edit Bin dialog box eleme
110. Element Description Panel anager The Panel Manager root node is the container for all panels It is the top level of the hierarchy Kits help users organize their panels in product or project specific folders Clicking a kit folder displays the Panel table o Erara marag E S AmpFISTR_ Panels _w1 H Blue v1 H E Green1 E f Frotiler_w1 The Panel folder icon represents a set of loci Clicking a panel folder displays the locus table For SNaPshot kits clicking a panel folder also displays a plot view Eg Panel Manager Elf AmpFiISTR_Panels_v1 Ef Blue v1 o po D351358 ae AALA das FGA The locus icon represents an individual locus Clicking a microsatellite locus displays the Bin view Clicking a SNP locus displays the Bin table GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands Panel Manager Window Commands Introduction The tables in this section describe the following Panel Manager views and menus Topic See Page File Menu 4 7 Edit Menu 4 11 Bins Menu 449 View Menu 4 19 File Menu The File menu offers you options to create or export kits duplicate import and export panels and import and export bin sets 7 F Panel Manager Le mime Edt Bins View Mew Panel Ctrl M Puplicate Panel Import Bin Set Ctrl Shift 6 Export Panels Ctri E Export Bin Set Ctrl Shitt E Export All Kite Ctri k New Kit Pan
111. G5S_F 6 Ir ID_Sarmpl Samples None Sample no expon HID_Advy Identifiler CE_G5_F ee Analyzing Samples i In the Samples view you have access to Sample Information Raw Data and EPT data Each of these tabs is discussed in the following pages GeneMapper ID Software v3 1 User Guide 3 5 Chapter 3 Using the Project Window Info Tab When a Sample file is selected from the navigation pane in the Samples view a special information window like that shown below is presented and Info is the active tab K l GeneMapper ID 3 1 Casework Project gmid Is Logged In File Edt Analysis View Tools Help e S E L i E Tate setin fio Tavie J05 8 3 Project E Casework H CO_Control fsa CO_Ladder fsa CO_Sample1 fsa CO_Sample2 fsa PP_Control fsa PP_Ladder fsa PP_Sarmple1 fsa PP_Sample2 fsa Progress Status info Raw Data EPT Data Sample Information Sample File CO_Ladder fsa Sample Origin Path D AppliedBiosystems GeneMapper Example Data HID Casework CO_Ladder fsa Status Message Analysis method lt HID_Classic gt modified Changed analysis method from HID Classic to Default Changed analysis method from Default to SNaPshot Default Changed analysis method from 5NaPshot Default to HID Classic File Source Disk media Error Message Message Current Settings Sample Type Allelic Ladder Pane
112. GeneMapper ID Software v3 1 User Guide B 29 Appendix B Software Genotyping Algorithms B 30 11eP2 11 Best Fit 2nd Order Curve AO 9 771623E 01 Al 1 220838E 01 A2 4 856885E 06 200 R 2 1 000 Size Calling Curve 2nd Order Least Squares I I I I I 1000 1500 2000 2500 3000 3500 4000 4500 Data Point Figure B 10 2nd Order Least Squares size calling curve Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper JD software v3 1 11eP2 11 Best Fit 3rd Order Curve AO 1 266506E 02 J Al 1 608974E 01 A2 2 041442E 05 A3 1 887158E 09 R 2 1 000 Size Calling Curve 3rd Order Least Squares I I 1 I I 1000 1500 2000 2500 3000 3500 4000 4500 Data Point Figure B 11 3rd Order Least Squares size calling curve Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper JD software v3 1 GeneMapper ID Software v3 1 User Guide Size Calling Methods for Classic and Advanced Modes Cubic Spline By definition the Cubic Spline method forces the sizing curve Interpolation through all the known points of the selected size standard Although Method _ this enforcement produces exact results for the values of the standards themselves it does not compensate for standar
113. Genotypes tables in the corresponding tabs Click OK to save the new setting Copying a Table To copy a Table Setting Setting l 4 ay Select Tools gt GeneMapper Manager to open the GeneMapper Manager window Note If the table is selected in the project window drop down menu then you can click the Table Settings icon or Tools gt Table Setting Editor Click the Table Settings tab Select a table setting name then click Open The selected setting opens Click Save As and give the table setting a new name Click OK to save the new table setting Editing a Table To edit a Table Profile 5 34 Setting l Select Tools gt GeneMapper Manager to open the GeneMapper Manager window Note Ifthe table is selected in the project window drop down menu then you can click the Table Settings icon or Tools gt Table Setting Editor Select the Table Settings tab Select a table setting name and click Open The selected setting opens Perform edits in the table setting Note See The Sample and Genotypes Tab Elements on page 5 32 Click OK when you have completed editing The changes you have made are saved to the Table Setting GeneMapper ID Software v3 1 User Guide Deleting a Table Setting Filtering Controls Section 5 3 Table Settings Tab To delete a Table Setting 1 Select the Table Settings tab in the GeneMapper Manager window 2 Select a table setting name and click Del
114. HID Genotyping E Panes fa e E aal E EA Pick SEAE E Izle X 323 67 Y 575 GeneMapper ID Software v3 1 User Guide Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued ltem Description Labels e Vertical Labels Labels are positioned vertically continued across the screen to prevent overlap This option is recommended for printing 5 x File Edit View Tools Alleles Help Plot Setting Hip Genotyping x Panes fa z EE ga Bi al E E fA E Pi E E E Sample Name Pand SQO SQ Samplel Profiler_Frus_v1 og cJ Desir D2zrsir DIsSs1 100 200 400 Samplel Profiler_Plus_vi jos pss Bsr Dss 0 100 201 300 400 xl gt Note Labels are enabled in Separate Dyes mode only for Microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot kit samples 8 12 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued Item Description Bins Ctrl shift B Controls whether or not transparent bins are shown or hidden on an electropherogram In the Binning mode bins are automatically displayed and the Bins menu item is checked Separate Dyes mode File Edit View Tools Alleles Help Plot Setting HID Genotyping z Panes
115. I 1 09s 1 0gm 1 0 3ppr 1 0 31pp 1 0spu 1 0an l 0gp d 0ce 1 0ov1 l los lgm 0 7pnr 0 71pu 1spu lan Ipp lcc lovu GQ 1 0 49 0 51 An overall GQ PQV of 0 51 displays a yellow flag Overall Size The thirteenth parameter tested in the Algorithm Testing category Quality PQV category is the final PQV for the overall size quality SQ Verification verification using both the Classic and Advanced modes 20 364 alleles for the overall size quality PQV were evaluated to test the overall sizing quality The sizing quality for each sample is used in the calculation of the overall GQ PQV Results The final size quality value for each sample displayed an accurate assessment of the sizing for each sample evaluated Example of Size Quality PQV Verification The samples view below is highlighting the sample for the Control DNA 9947A and is displaying a red size quality value As shown in Figure I 23 on page I 35 the start scan collection point entered in the analysis method excluded the 75 base pair size fragment peak resulting in the inaccurate labeling of the size standard peaks GeneMapper ID Software v3 1 User Guide I 33 Appendix GeneMapper ID Software Verification GeneMapper ID 3 1 Database gmid Is Logged In S ENE gt A Ho Table jma all E E Project E Ef Databasing a ater SS a Ce Ce oe Sa a 1 Identifiler Identifiler_v1 Databasing ABI3100 lt a ee 9 Adv
116. Increasing the Polynomial Degree Value Optimizing Peak Detection Sensitivity Example 2 The figure below shows the data presented in the figure above see page B 22 re analyzed with a window size value of 10 and polynomial degree value of 5 s 1820 1830 1840 1850 1860 1870 1880 1890 1900 1910 1920 1930 1940 1950 1960 1970 1980 1990 2000 2010 2020 2030 2040 p O OS S O A O A O O O S S S S Cs os ie en O O N O A es oa 1800 1600 Figure B 6 Electropherogram showing all four peaks detected after reducing the window size value and increasing the polynomial degree value GeneMapper ID Software v3 1 User Guide B 23 Appendix B Software Genotyping Algorithms Optimizing Peak Detection Sensitivity Example 3 Effects of The figure below shows the result of an analysis using a peak Extreme Settings window size value set to 10 and a polynomial degree set to 9 This extreme setting for peak detection led to several peaks being split and detected as two separate peaks a cn A a aa aa ee fl oll Figure B 7 Electropherogram showing the result of an analysis using extreme settings for peak detection B 24 GeneMapper ID Software v3 1 User Guide Peak Detection Slope Threshold for Peak Start and Slope Threshold for Peak End Parameters Peak Detection Slope Threshold for Peak Start and Slope Threshold for Peak End Parameters About These Parameters How These Parameters Work Guidelines for Using These Parameter
117. M 377 to a sample or gel file made on another ABI PRISM 377 In other words you cannot apply matrix created on one instrument to other instruments of the same model You can only assign a matrix to sample files generated on the same instrument under the same electrophoresis gel matrix and buffer conditions and using the same dye set GeneMapper ID Software v3 1 User Guide 5 55 Chapter 5 Using GeneMapper Manager When to Create a Create a new matrix in the following conditions New Matrix For each dye set F and G5 e When you change the dye set you use to label sample fragments for example if you are using the fifth dye e When you see multiple unexpected peaks of different colors under an expected peak e Anytime a service engineer changes an instrument part performs preventive maintenance or realigns the optics For information on when to create a new matrix refer to Protocols for Processing AmpFtSTR PCR Amplification Kit Products with the Microsoft Windows NT Operating System PN 4341742 Where to Store Store exported matrix files mtx intended for use by Data Collection Exported Matrix software in Files D AppliedBio Shared Analysis Sizecaller Matrix If Data Collection and Analysis are installed on different computers the location is the same Remember to copy the matrix from the analysis computer to the Data Collection computer Note GeneMapper ID software matrices are stored in the Oracle data
118. Manager window If there are multiple bin sets associated with a kit use the drop down menu to select the appropriate bin set to be deleted Note This action cannot be undone Show Project Alleles The Show Project Alleles command displays all of the marker s allele calls that are present in the genotypes table of the open project Allele calls are displayed as blue asterisks This option is enabled when a marker is selected in the Panel Manager navigation pane and a profile exists for that marker The option is for microsatellite data only Add Reference Data The Add Reference Data command opens the Add Reference Data dialog box which allows you to associate data with a microsatellite panel or SNaPshot kit for creating markers and bins For microsatellite panel reference data samples must have been analyzed using the panel for which the data serves as a reference and the appropriate size standard For SNaPshot kit reference data samples must have been analyzed using the appropriate size standard and analysis method with SNP genotyping as the analysis type GeneMapper ID Software v3 1 User Guide 4 17 Chapter 4 Using Panel Manager Panel Reference Data This feature works for both SNP genotyping and microsatellite data e SNP genotyping data The Panel SNP Reference Data command opens the SNP Reference Data dialog box which allows you to organize the reference data imported at the kit level into panels This feature is
119. Mapper ID Software v3 1 User Guide B 17 Appendix B Software Genotyping Algorithms Guidelines for Using These Parameters Examining Peak Definitions Effects of Varying the Polynomial Degree To detect well isolated baseline resolved peaks use polynomial degree values of 2 or 3 For finer control use a degree value of 4 or greater As a guideline set the peak window size in data points to be about 1 to 2 times the full width at half maximum height of the peaks that you want to detect To examine how GeneMapper JD software has defined a peak select View gt Show Peak Positions The peak positions including the beginning apex and end of each peak are tick marked in the electropherogram The figure below depicts peaks detected with a window size of 15 data points and a polynomial curve of degree 2 green 3 red and 4 black The diamonds represent a detected peak using the respective polynomial curves Note that the smaller trailing peak is not detected using a degree of 2 green As the peak detection window is applied to each data point across the displayed region a polynomial curve of degree 2 could not be fitted to the underlying data to detect its structure Polynomial curve of degree 4 black Polynomial curve of degree 3 red Polynomial curve of degree 2 green f Figure B 1 Electropherogram showing peaks detected with three different polynomial degrees GeneMapper ID Software v3
120. PO 100 110 4120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 230 300 310 320 330 340 350 ee a S S S S ES Sample3 Identifile_v1 tt tt tt tt ttt tt tt tt D351358 Riis 0 1 eee 100 M0 120 130 140 450 160 170 180 190 200 800 D13 317 D16 539 D2 1338 240 220 230 240 250 260 270 280 290 300 310 320 330 340 350 ter iri tr toar sk et ID19S433 TPOX 18551 FWA D 100 110 4120 130 440 150 160 170 180 1490 200 240 220 230 240 250 260 270 280 290 300 310 320 330 340 350 Sampled Ideia v 4 o A D55818 FGA 100 110 120 130 440 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 Fie Edit Yiew Tools Alleles Help a x Note Bins are enabled in Separate Dyes mode only for microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot kit samples When you turn off a specific dye color for SNaPshot kit samples the bins for that color turn off also Peak Shows hides the peak position indicators on the Positions electropherogram currently viewed Checked when active Disabled for overlaid samples 0 x Plot Setting HID_Genotyping_display x Panes f z ECT EEPE fi VEN Vic E e EE Sample Name Pand Marka OS BIN PHR LPH SPU AN BD CC OVL GQ ere are no controls in this selectii Sample Identifile_v1 Dss C O E O C C O C Aj 400 300 100
121. Plot window Move the mouse over each icon to view the tooltip description The tooltip describes what action occurs if the button is pressed For example by default all colors are shown If you move your mouse over the All icon the tooltip says Hide All Dyes If you click the icon all colors will be hidden GeneMapper ID Software v3 1 User Guide 5 51 Chapter 5 Using GeneMapper Manager 5 52 GeneMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix section 5 5 Creating and Evaluating a Overview Matrices Tab Elements Matrix This chapter applies only to the ABI PRISM 310 Genetic Analyzer and the ABI PRISM 377 DNA Sequencer instruments This section describes the process of creating and evaluating a matrix With the GeneMapper I D Software Version 3 1 matrices are stored in the GeneMapper JD database They become mtx files on your hard drive only when you export them You must export a file in order to run data collection software on the ABI PRISM 377 DNA Sequencer You have the option of selecting four or five dyes depending on the application when creating a new matrix for data collection Table 5 14 Description of elements in the Matrices tab ltem Description Name column Matrix name Last Saved Date Time stamp showing when the matrix was column last saved Owner column User name of the person who created the matrix of Dyes column The number of dyes s
122. Project and Save boxes enable you to save projects to the Save Dialog database The first dialog box is presented for the Save command if Boxes you want to save a project before it is analyzed Using Save after analysis does not require a dialog box because you are required to name a Project before analysis im Save Project x Q Project name canos The Save dialog box opens when you select the Save As command GeneMapper ID Software v3 1 User Guide C 29 Appendix C Project Window Software Interface Save Project name FEWE Dont Save Cancel The elements of these dialog boxes are described in the table below Element Description Project name Text field for the name of the project accepts alphanumeric characters The database requires standard Windows interface restrictions on length and symbol characters OK Saves the project to the database Saved projects are tagged with the Project Name User Name and Time Date Stamp If the name you entered already exists an alert message is displayed Cancel closes the alert dialog and displays the Save Project dialog box again Cancel Closes the dialog box and does not save the project Save Saves the project under the name entered in the dialog box Don t Save Aborts the Save action C 30 GeneMapper ID Software v3 1 User Guide Project Window File Menu Dialog Boxes Export Table The Export Table dialog box allows y
123. SFIPO G E 7 TRUE FALSE ol 2 2 of of of of 2 olos 15 COfiler vl D75620 Y G 7 TRUE FALSE ol 2 of of of ol 2 oogs 16 Pro ler_PI 0351358 E 14 15 FALSE FALSE 0 0 0 OF 00 00 o0 05 Figure 9 1 Example of an exported table with PQVs displayed in an Excel spreadsheet It is recommended that you create a separate table setting to use just for exporting if you do not want to have all of these PQVs within the exported table Or you can manipulate the table once it is imported into a spreadsheet to remove these columns To export a table 1 Click the tab of the table you want to export Samples or Genotypes 2 Select File gt Export Table GeneMapper ID Software v3 1 User Guide 9 3 Chapter 9 Exporting Table Data 3 Complete the Export Table dialog box Export Table f a x Look in GeneMapper f c Export File 4s Tab delimited text txt _ Plot Settings Tab delimited text txt _ Size Standards Comma separated values csv __ Table Settings My Documents File name casework_1 Export Table ee Files of type frav and Comma delimited Text txt csv bi Cancel P Navigate to the location you wish to export the file o Select to export the file as Tab delimited text txt or Comma separated values csv from the Export File as drop down list O Type in the desired File Name The default file name is the name of the project d Click Export T
124. The Delete all overlapping bins and out of bounds bins option shown below does not affect the marker allele size range all overlapping and out of bounds bins are deleted 3 Delete all overlapping bins and out of bounds bins A B C D EF G cCc D E Marker Range 100 110 Marker Range 100 110 Note If all the bins in an import are inside their marker ranges the dialog box does not open Export All Kits The only way to back up panels and bin sets is to export them You will be prompted for a location and name for the exported item The Export All Kits command opens the Export All Kits dialog to allow exporting all kits and their associated panels markers and bin sets to a selected location When you use this option panels retain their current panel name as listed in the Panel Manger Bins are named as follows Kit name Bin set name _bins txt Edit Menu The Edit menu contains the Undo command and the Clear command sali Bins view Undo The undo command allows for the Clear command to be undone GeneMapper ID Software v3 1 User Guide 4 11 Chapter 4 Using Panel Manager Clear The Clear command deletes items selected in the navigation pane from the GeneMapper JD database e Inthe Kits file menu Clear deletes the kit and all bin sets associated with that kit e In the Panels file menu Clear deletes the panels In the Markers file menu Clear deletes the markers Use the Bins Menu items to delete bins a
125. Up Data Transferring Data Logging into Another GeneMapper Database from a Networked Computer Transferring Data from One Copy of GeneMapper to Another Exporting Importing F 2 If you want to view re analyze a project created on a different copy of GeneMapper computer you have two option e Login to the GeneMapper database where the project is saved from a networked computer with GeneMapper installed Refer to Chapter 2 New Host Option for more details on logging into a host computer e Export the project from GeneMapper Manager Projects Tab window on the computer it was created on and import it into the GeneMapper Manager on the computer you wish to view it on IMPORTANT If you want to open a project that was created on a different computer by exporting and importing there are several things you must keep in mind and several components linked to the project that need to be in place in order to have full access to the information contained within the project Therefore follow the instructions below to ensure proper transfer of data This will allow you have full access to the project You will be able to re analyze the sample files apply new analysis settings view all plots raw data EPT data genotypes table and allele calls However you will not be able to save this project in your database If you want this project saved in your database you must export it and import it into your database manually To v
126. _vl oOYss820 6 z 11 sample4 Databasing Identifiler_yvl CSF1FP 6 10 13 Sampled Databasing Identifiler_vl 0351358 G 14 1 Sampled Databasing Identifiler_vl THL G G 9 3 sample4 Databasing Identifiler_vl D135317 G 12 13 sample4 Databasing Identifiler_vl D16553 G 12 13 sample4 Databasing Identifiler_vl D251338 G 25 Sampled Databasing Identifiler_vl o195433 Y 12 14 sample4 Databasing Identifiler_ vli vwa Y 16 18 Sampled Databasing Identifiler_vl TPox Y z sample4 Databasing Identifiler_vl D18551 18 Sampled Databasing Identifiler_ w1 AMEL F sample4 Databasing Identifiler_vl D55818 FR 12 sample4 Databasing Identifiler wl FGA F z1 26 Samples Databasing Identifiler_vl O851179 6 12 13 Samples Databasing Identifiler_vl D21511 6 29 30 4 Ha Note Additional pages are created to accommodate the number of rows in the table Each additional page that continues the table to the right would have the same row numbers as shown in the left column GeneMapper ID Software v3 1 User Guide C 35 Appendix C Project Window Software Interface Example Plot Printout Samples Plot AE Fie Edit view Tools Alleles Help Plat Setting HID Genotyping E Panes fa E E aal E r E ha fle AO LS Sample Name Pand SQO SQ E xX 217 69 Y 163 Bin 5 Marker TPOX Note The number of plots per page is set by the Panes dialog box in the Plots window The example above was scaled down vertically and would be prin
127. a The same single source DNA sample was amplified with the AmpF STR Identifiler PCR Amplification kit processed on the 310 for Macintosh and NT OS then analyzed with different versions of software to compare GeneScan for the Macintosh OS to the Classic mode in GeneMapper JD and to compare GeneScan for Windows NT OS to the Advanced mode in GeneMapper ID software Note the similarities in peak heights measured in RFUs Table l 5 Peak height example data analyzed using different software Software and Version To view the data see GeneScan Software v3 1 2 Figure l 1 on page I 10 Macintosh OS and Genotyper Software v2 5 2 GeneMapper D Software v3 1 in Figure l 2 on page I 11 the Classic mode GeneScan Software v3 7 1 Figure l 3 on page I 12 Windows NT OS and Genotyper v3 7 GeneMapper ID Software v3 1 in Figure l 4 on page I 13 the Advanced mode GeneMapper ID Software v3 1 User Guide l 9 Appendix GeneMapper ID Software Verification 0 sss p63 enti SSS Fy 110 120 130 140 150 160 170 180 190 z200 210 220 230 240 250 z260 270 280 80 0010 z320 S040 110 iz z0 130 i 0 150 160 i70 180 190 z200 710 220 SO 240 250 z260 FO 280 290 300 310 z320 330 340 E 44 Female_a2_ing 4Blue Female_a2_ing z 1500 i 1000 E so S g H 10l 10l 1582 15859 1098 1353 1082 968 1500 11 44 Female 52 ing 4 Green Female 5z2 ing 2000 1000 H 11 11 1756 2234 1972 1708 1733 1573 44 Fema
128. a analysis at any time to display information in a different table format Analyzing the To analyze the data Data l Click Analyze F GeneMapper ID 3 1 test gmid Is Logged In File Edit Analysis View Tools Help ek ce S A A e Hl J Project Casework Table Settings it Samples Genotypes Note The peak detection algorithm Classic or Advanced selected in the Analysis Method must be the same analysis method used to create the size standard definition The pre defined size standard definitions that are supplied with the software were created using the Basic Advanced peak detection GeneMapper ID Software v3 1 User Guide 7 9 Chapter 7 Analyzing and Interpreting Data 10 algorithm If you want to use the Classic peak detection algorithm define a custom classic size standard definition If the analysis method and size standard do not match the software issues the following alert when you press the analyze button SS LLtCS There are samplets that do not meet analysis requirements Please see Error Message in the Info view of each sample Do vou want bo continues 2 At the Project name prompt type in the desired name of the project in the Save window then click Save Project name Save Dont Save Cancel During analysis you can see the progress along the bottom of the project window Once completed it will read Analysis Completed EEE I
129. abase Sequence aiena Samples To Add r Search Criteti Condition Criteria Project Project Name starts with Sample Project Create Date is Analysis Genotypes Project Modification Date lis Search Results cancel Search Add To List gt gt Hesr z Fide Analyze Cancel Preferences This feature allows mixing and matching sample files For example you can merge sample files from a number of small projects into a single large project C 42 GeneMapper ID Software v3 1 User Guide Add Samples to Project Dialog Criteria and Rules Description Search criteria This list and table contains the search criteria from which to extract samples from the GeneMapper D database Note Locations of the list and table are pointed out in the figure on the previous page e Criteria selector list list of properties columns in the project window grouped by topic e Field column Text field that contains the properties columns in the selected group criterion e Condition column Combo box that contains conditions that the system will use to search for samples Search rules include ls Contains Starts With Date criteria Presents a calendar widget to specify the date e Criteria column Field for specifying the text string associated with the search rules which are case sensitive Rules
130. able Opening Note You need to adjust column widths in Microsoft Excel even if Exported you adjusted them appropriately in the GeneMapper JD software Samples or E Project Tables in If you exported the files as a Tab delimited text txt file Microsoft Excel 1 Find the exported table file on your hard drive 2 Right click on the file and select Open With gt Choose Program 3 From the list of applications choose Microsoft Excel for Windows The table opens within a spreadsheet If you exported the files as a Comma separated values csv 1 Find the exported table file on your hard drive 2 Double click to open the file The table opens within a spreadsheet 9 4 GeneMapper ID Software v3 1 User Guide About CODIS About CODIS GeneMapper ID GeneMapper ID software can export data from the analysis in a Software format suitable for the FBI Laboratory Combined DNA Index Features System CODIS For more information about CODIS see http www fbi gov hq lab codis index1 htm CODIS Creation of CODIS CMF files from GeneMapper JD software Requirements requires that e A specimen category is selected for each sample to be exported e Samples designated as a positive control negative control or allelic ladder cannot be exported e Allele calls contain acceptable characters numbers decimal points letters X and Y for Amelogenin allele calls and the symbols less than lt and greater than gt GeneMapper
131. able is customizable and can be easily exported e Genotypes View See page C 7 The tab for this view is enabled after successful analysis Samples that do not have a Panel attribute do not display in the Genotypes table GeneMapper ID Software v3 1 User Guide 3 11 Chapter 3 Using the Project Window Navigating in the Project Window Navigation in Samples and Genotypes Views This section provides information on how to navigate as well as how to access different views for samples files e Navigation in Samples and Genotypes View e Info Tab See page 3 6 e Raw Data Tab See page 3 7 EPT Tab See page 3 8 The left panel of the Project window has a tree view navigation device In the Samples view the tree viewer has three levels Each sublevel of the tree view is indented to the right In the Genotypes view the tree viewer has three levels and each sublevel of the tree view is again indented to the right Samples View Genotypes View Project e Panels root level Run folder e Panels Sample Markers loci As soon as you click the tree view controller indicated in the figure below the tree view expands with the subordinate level indented Clicking the controller a second time collapses the level back to its original form E Project Project Casework Run folder pe cO Control fsa CO_Ladder tsa a CO Sample tsa vee CO Sample tsa Sampl
132. agment and another value is assigned 4 The two size values are averaged to determine the unknown fragment length GeneMapper ID Software v3 1 User Guide B 33 Appendix B Software Genotyping Algorithms Global Southern This method is similar to the Least Squares method in that it Method compensates for standard fragments that may run anomalously The method creates a best fit line through all the available points and then uses values found on that line to calculate the fragment values 11 P27 11 Best Fit 2nd Order Curve AO 9 7716235E 01 Al 4 1 220838E 01 AZ 4 856065E 06 R 2 1 000 Size Calling Curve Global Southern Method I I I I I I I I 1000 1500 2000 2500 3000 3500 4000 4500 Data Point Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper ID software v3 1 The Equations The following table describes how the equations work Equation Description L c m m0 LO Attempts to describe the reciprocal relationship between the mobility m and the length LO of the standard fragments Ddi Li c mi m0 LO 2 The fitting constants LO mO and c are calculated by a least squares fit to minimize the left side quantity B 34 GeneMapper ID Software v3 1 User Guide Size Calling Methods for Classic and Advanced Modes How This Method Works All points in the standard are weighted
133. aluating a Matrix on page 5 53 Smoothing Used to help reduce the number of false peaks detected by the GeneMapper ID software You have the following options Select To None apply no smoothing Select this option if the data has very sharp narrow peaks of interest Light provide the best results for typical data Heavy apply to data from slower runs that has very broad peaks or to avoid the detection of sharp edges Choosing this option might reduce peak size or eliminate narrow peaks GeneMapper ID Software v3 1 User Guide Classic Mode Peak Detection Algorithm Settings Size Calling Click a radio button to select the desired size calling method The Method GeneMapper JD software uses these methods to determine the Parameter molecular length of an unknown fragment Options Description of the size calling method parameter options Item Description 2nd Order Least Squares and 3rd Order Least Squares Both Least Squares methods use regression analysis to build a best fit size calling curve Cubic Spline Interpolation Forces the sizing curve through all the known points of the selected GeneScan size standard Local Southern method Determines the sizes of fragments by using the reciprocal relationship between fragment length and mobility Default method Global Southern method Similar to the Least Squares method in that it compensates fo
134. an item to view its description See also My Documents My Network Places My Computer f objectis i bytes My Computer c Transfer the original sample files fsa files generated by Data Collection software into the appropriate folder in this case it is the Casework folder w Casework File Edit wiew Favorites Tools Help da Back p G Search Oh Folders 4History i CO Control Meco Ladder us jco Sample Casework Mco Sample APP Control Pod PP_Ladder description BPP Samplet BOJPF_Samplez Select an item to wiew its See alsa My Documents My Network Places My Computer 8 objectis 742 KB My Computer GeneMapper ID Software v3 1 User Guide Appendix F Transferring Data and Backing Up Data To view raw data EPT data and re analyze imported projects without establishing the link 1 Make sure the GeneMapper database you are importing the project into contains the same exact panels bins analysis method and size standard originally used to create the project If this is not the case export these items from the original copy of GeneMapper and import them into the GeneMapper copy you would like to view the data on 2 Import the sample files fsa files into a blank project 3 Select the appropriate analysis settings and analyze the project F 6 GeneMapper ID Software v3 1 User Guide Backing Up Data and Cleaning the Database Backing Up Data and Cleaning the Database Backing U
135. anced eis ase avo Sa fee aO lt a _ so Advanced pe phases o a a a a i Bo o O eee ee Figure l 22 The sample table displaying a red size quality value I 34 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Size Match Editor Ee l File Edit view Tools ID Control tsa Size Matches Size Calling Curve Sizing Quality 0 0 200 2890 Mis labelled size standard peaks iI Boog Food ggi Ok Cancel pplyr Figure I 23 Size Match Editor in the Advanced mode displaying the size standard for the control DNA 9947a amplified with the AmpF STR Identifiler kit displaying a size quality value of 0 0 GeneMapper ID Software v3 1 User Guide I 35 App endix GeneMapper ID Software Verification Workflow Test Parameters Process Samples Per Color Per Sample and Per Marker Display Default and Customized I 36 Plot Settings Printing The third category presented is Workflow testing 1 Sort and display samples per color per sample and per locus functionality test 2 Display default and customized plot settings functionality test 3 Printing test functionality test The first parameter tested in the Workflow category is the ability to sort and display samples per color per sample and per locus The genotype plot view enables a user to graphically sort and display data per sample per color and per locus and to assess the quality of
136. are This chapter describes hardware requirements and procedures for installing registering and logging in and out of the GeneMapper ID software v3 1 This chapter covers Hardware Requirements 04 244 sande244c0iu bes daboiadewsss 2 2 Installing the GeneMapper JD Software 0000 2 3 Registering GeneMapper JD Software 0 000000 2 6 Logging Onto and Logging Out of the GeneMapper JD Software 2 7 GeneMapper ID Software v3 1 User Guide 2 1 Chapter2 Getting Started with GeneMapper ID Software Hardware Requirements Hardware The following table describes the components your computer system requires to run GeneMapper JD software Requirements System Component Minimum Requirement Recommended 17 inch monitor Computer e Intel Pentium Ill processor Intel Pentium IIl processor faster minimum of 540 MHz than 700 MHz e 256 MB RAM e 512 MB RAM e 6 GB hard disk free space e 10GB EIDE hard drive 6 GB free space e 20 48X IDE CD ROM e 10 100 NIC with RWV internal Monitor 800 x 600 pixels size 1024 x 768 pixels size 19 inch or larger monitor Operating System Microsoft Windows NT v 4 Service Pack 5 0 Microsoft Windows 2000 Professional Service Pack 3 or Windows NT v 4 0 Service Pack 5 Ethernet Capability Network card for Oracle installation TCP IP must be installed prior to Oracle installation Network card for Oracle
137. are and software be restored to the standard configuration prior to providing service or technical support For systems that have built in computers or processing units installing unauthorized hardware or software may void the Warranty or Service Plan Limited Product Warranty Limited Warranty Applied Biosystems warrants that for a period of ninety 90 days from the date the warranty period begins its GeneMapper ID Software Version 3 1 will perform substantially in accordance with the functions and features described in its accompanying documentation when properly installed on the instrument system for which it is designated and that for a period of ninety 90 days from the date the warranty period begins the tapes diskettes or other media bearing the software product will be free of defects in materials and workmanship under normal use If buyer believes that it has discovered a failure of the software to satisfy the foregoing warranty and if buyer notifies Applied Biosystems of such failure in writing during the ninety 90 day warranty period and if Applied Biosystems is able to reliably reproduce such failure then Applied Biosystems at its sole option will either 1 provide any software corrections or bug fixes of the identified failure if and when they become commercially available to buyer free of charge or 11 notify buyer that Applied Biosystems will accept a return of the software from the buyer and upon su
138. atabasing Identifiler_vl 06351358 G ID_Neg_cntr Databasing Identifiler_vl THL G ID_Neg_cntr Databasing Identifiler_vl D135317 G ID_Neg_cntr Databasing Identifiler_ w1 D16553 G ID_Neg_cntr Databasing Identifiler_vl D251338 G ID_Neg_cntr Databasing Identifiler_vl D195433 Y ID_Neg_cntr Databasing Identifiler wi wwa Y ID_Neg_cntr Databasing Identifiler wi TPO Y ID_Neg_cntr Databasing Identifiler_ w1 D18551 ID_Neg_cntr Databasing Identifiler_ w1 AMEL F ID_Neg_cntr Databasing Identifiler_vl D55818 R ID_Neg_cntr Databasin Tdentifiler_vl FGA F Samples Databasing Identifiler_vl D8S1179 6 12 14 Samples Databasing Identifiler wil D21511 6 30 31 Samples Databasing Identifiler_vl D75820 6B G 10 Samples Databasing Identifiler_yvl CSF1P 6 10 12 Samples Databasing Identifiler_vl 0351358 G 15 18 Samples Databasing Identifiler_vl THOL G 7 z Samples Databasing Identifiler_vl D135317 G G 13 Samples Databasing Identifiler_vl D16553 G G 11 Samples Databasing Identifiler_vl 0251338 G 22 Samples Databasing Identifiler_vl o19s433 Y 12 15 Samples Databasing Identifilervl vwa Y 15 17 Samples Databasing Identifiler_vl TPO Y z 12 Samples Databasing Identifiler_vl D18551 15 18 Samples Databasing Identifiler_ w1 AMEL F Samples Databasing Identifiler_vl D55818 FR 12 13 Samples Databasing Identifiler_vl FGA F 24 28 Sampled Databasing Identifiler_vl D8S1179 6 10 16 Sampled Databasing Identifiler_vl D21511 6 28 29 Sampled Databasing Identifiler
139. ated based on peak heights of the called allele peaks e For SNP the ratios are calculated the same as for microsatellite markers except it goes across two different colors and only two peaks are used in the calculation e Not editable GeneMapper ID Software v3 1 User Guide Using Process Component Based Quality Values The PQV flags on the Genotypes page continued Column Description LPH Low Peak Height e A flag Pass Ii or Check A is automatically displayed when the alleles are lower that the specified values and do not result in the proper intensity The homozygous default is 200 and heterozygous default is 100 values can be set in the Peak Quality tab of the Analysis method Analysis Manager Setting this flag reduces the final PQV Genotype value GQ parameter The default is 50 0 5 multiplier of what it would be otherwise Not editable SPU Spectral Pull Up e A flag Pass Il or Check A is automatically displayed when the marker signal contains bleed through peaks pull up peaks Pull up is when the peak height of the called allele peak is less than X of the larger peak within 1 data point The default is a conservative 10 to check all colors The user can set this value This flag is used for HID SNP and Microsatellite Not editable BD Broad Peak A flag Pass Ii or Check A is automatically displayed when the called alleles peak width is wide
140. ated menu items as described throughout this chapter Position your mouse over an icon to view its tooltip description 1 2 3 4 5 6 8 EKO ERIN E ense arme gt Ewe S E Table 4 1 Panel Manager icons and descriptions Item Description 1 z New Kit Panel Locus icon automatically pet updates to reflect the current action depending on the item selected in the navigation pane File menu 2 Clear icon performs kit panel locus x deletion Edit menu 3 mea Icons select bin options Add Bin Delete Bin Edit Bin Bins menu 4 ry Icon allows viewing of Project alleles beck Bins menu 5 rT Icon switches view to full X Axis and Y Axis scales View menu 6 Ese anarsa Bs E Pull down menu allows selection of a lll eels bin set for a selected kit 7 Icons select specific kit panel locus tions Bins menu Me ES e E New Bin Set lll Add Reference Data Auto Bin Ei Panel Reference Data E Auto Panel 4 8 Dye color toggle icons View menu pmm mm f om ie fe GeneMapper ID Software v3 1 User Guide 4 5 Chapter 4 Using Panel Manager Panel Manager Navigation Pane 4 6 The Panel Manager navigation pane is the primary navigation interface in the Panel Manager window Since it is found in all Panel Manager views as shown in the table below it enables moving back and forth between the views to view edit and create kit folders panels locus and bins The elements of this field are described below
141. atio Low Peak Height Spectral Pull up Allele Number Broad Peak Double Peak SNP Narrow Bin SNP Control Concordance Overlap HID Show Pass green squares or Show Check yellow triangles Show All Records Show All Records show Pass show Check Allele Edit Allele Display Overflow Yes or No Show All Recards Show All ecards Genotype Quality GQ Show Pass Show Check or Show Low Quality Show All Rec hal Show All Records show Pass Show Check Show Low auality GeneMapper ID Software v3 1 User Guide 5 39 Chapter 5 Using GeneMapper Manager 5 40 GeneMapper ID Software v3 1 User Guide Section 5 4 Overview Plot Settings Tab Table 5 12 Description of elements in the Plot Settings Tab Elements Section 5 4 Plot Settings Tab Plot Settings Tab The Plot Settings tab enables you to create and edit plot setting profiles for viewing your data in the Samples and Genotypes plot views This section describes how to use the Plot Settings tab in the GeneMapper Manager window of the GeneMapper ID Software Version 3 1 and how to use the features of the Plot Settings Editor To display the Plot Settings Tab select Tools gt GeneMapper Manager gt Plot Settings F GeneMapper Manager Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Name Last saved Owner Description Sizing 2003 06 06 16 02 22 grid Fact
142. ats of Panel and Bin Text Files Bins Definition The examples below show bins defined in the format used by Example GeneMapper ID Software v3 1 in a tab delimited text file txt using notepad or wordpad or an Excel spreadsheet saved as a txt file Note The allele size range ASR in the bin definitions file should match the ASR in the Panels definition file Also kit panel and marker names must be the same between the two files fj AmpFLSTR_Bins_v 1 Notepad File Edit Format Help GenemMapper ID v3 1 Last edited 071403 GMv3 0 For import into GenemMapper ID v3 1 GMy3 0 AMPFLSTR_Panels_v1 AMPFLSTR_Bins_v1 5 6 Blue_v1 7 8 9 pei a Se a a a a a a w a a aa r WU uo an an an an on nin Le a a a a a a O Wu oo nann WU UU UU on on en en in an an an Lae O bO a a a a a a DODO DO D0 WU oo nin G Wun on uA anan in anan in Vv Q 0 Q Os Q Q Q Q Q QO Q Q Q Q QO F Q QO QO 0 Q Q Q QO 0 Q QO e a a a a a a a a a WUT UU on on on on nin 1 2 3 4 Note See column and row descriptions on the following page GeneMapper ID Software v3 1 User Guide 4 27 Chapter 4 Using Panel Manager Column Description 1 Bin name The name of the allele at an expected location within a locus 2 Bin center The location of the bin center in base pairs e g 110 0 5 bp 3 Bin left offset The left boundary of the bin expressed as
143. ayed at the apex and base may be difficult to visualize of this D21S11 allele Spectral pull up is measured for the VIC dye labelled peak in the THO1 size range The off scale flag yellow and the spectral pull up flag green for D21S11 are correct However the off scale flag yellow and the spectral pull up flag green for THO1 are incorrect GeneMapper ID Software v3 1 User Guide Results of Verification Testing Peak Height Ratio and Allele Number PQV Flag Verification The eighth and ninth parameters tested in the Algorithm Testing category were the individual PQV for the peak height ratio PHR flag verification and allele number AN flag verification using both the Classic and Advanced modes 3 678 alleles from mixture DNA samples and 16 alleles from a single source DNA sample for the peak height ratio flag and allele number flag were evaluated Results 100 of the alleles tested accurately reflect the PHR and AN PQV Example Data Showing PHR and AN PQV Flag Verification GeneMapper ID 3 1 Database gmid Is Logged In O x File Edit Analysis View Tools Help e S S e L EL GB Tavie Setting Hi0 Table H 8S GS Panels Samples Genotypes Aldent Sample Name Run Name Panel Marker Dye Allele1 Allele2 ADO AE OS BIN PHR LPH SPU JAN BD CC OYL GQ 69 Sample4 Databasing Identifiler_v1 D3513568 G 14 17 a z E m s z E 70 Sample4 Da
144. base 5 56 GeneMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix Process of Creating a New Matrix The following figure shows the procedure for making a new matrix Import samples into a new GeneMapper D project View raw data to determine data point range for the matrix calculation and sample quality Insert the data point to start at for each dye color Click Create Review and edit the matrix result Click OK to save and close the window Figure 5 29 Creating a New Matrix Project window GeneMapper Manager window Matrix Maker window For sample preparation and loading information refer to the appropriate instrument user guide GeneMapper ID Software v3 1 User Guide 5 57 Chapter 5 Using GeneMapper Manager Steps to Create a Table 5 15 Steps to create a new matrix New Matrix Step Process See Page 1 Choosing a Data Point Range 5 59 2 Generating a New Matrix 5 60 3 Assigning the Matrix to Samples 5 63 5 Using the Matrix with Data Collection 5 63 6 Causes for Bad Matrices 5 64 Choosing a Data Point Range for the Matrix Calculation Introduction Viewing the Raw 5 58 Data Depending on how well your Matrix Standards run it may be necessary for you to choose a specific range of data points to be considered for your matrix calculation In order to choose appropriate values for the data point range you must first view the sample
145. ble view In the Panel Table view you can e Edit panel names and enter comments in the panel table e Create new panels using the New Panel command e Export panels and bin sets used for project analysis to backup Note Editing or deleting kits panels and bin sets prevents you from displaying the original data Export any panels and bin sets that you might use at a later date or for reanalysis of old projects ig Panel Manager Kit Mare Kit Type AmpFLSTR_Panels_ 1 AmpFLSTR Panels w1 Microsatellite Reference Samples OK Cancel Apply Note A warning alert is shown whenever kits or panels are deleted to remind you that this reference data may have been used previously in sample analysis 4 34 GeneMapper ID Software v3 1 User Guide Panel Table View Panel Table The table below lists the columns in the Panel table Col umns LETE Manager Fie Edit Bins wiew E E En se ampFLSTR_Bins_v ih Eee es Ho a Comment suk X FF ES Panel Manager E MICROSATELLITE Y Green Iwi none H E AmpFLSTR Paneis v1 Profiler_v1 none Profiler_Plus_CODIS_v1 none COfiler_CODS v none Identifiler_CODIS_v1 none Note All columns are resizable and editable Column Description Name p Panel Contains the name of the panel Cell accepts alphanumeric Name characters restricted for any invalid symbol characters includes all Microsoft Windows invalid characters Note When
146. cally calling alleles or performing genotyping F GeneMapper ID v3 1 Untitled gmid Is Logged In l l x File Edit Analysis View Tools Help E gge elm Be a E E E eve setting ampriste Taie E E Project Samples Senatypes Status Sample File Sample Name Sample ID Comments Sample Type Specimen Cater Analysis Method Panel Size Standard 1 2 3 4 i 7 8 E 10 Analysis Completed Stop For more detailed information on the main project window and its functions and features see Appendix C Project Window Software Interface 3 2 GeneMapper ID Software v3 1 User Guide Overview of the Project Window The Project window makes it easy for you to e Add samples initiate analysis and export results e View sample data and access data plots associated with a particular sample file The pre analysis raw data version of the data in the Samples view of the Project window The tabular results of analysis in the Genotypes view of the Project window or Plot s of selected Project sample data in the Plot window e Initiate the process of creating analysis methods and importing and defining panels and bins Overview of the Project Window Project Window _ In the Project window you can Interface 3 e Minimize the window to an icon on the Microsoft Windows Conventions
147. ce ltem Description Enabling Show Offsets Displays bin offsets for Always enabled selected samples Options Opens the Options dialog Always enabled box See Options Dialog Box on page C 18 Options Dialog Box The Options dialog box contains several tabs for setting preferences associated with various software features The following tabs are described below Topic See Page Startup Tab C 19 Add Samples Tab C 20 Analysis Tab C 22 Users Tab C 24 Note Each registered user has his or her own set of preferences When you set these options it only affects the currently logged in user C 18 GeneMapper ID Software v3 1 User Guide Startup Tab Project Window Menus The Startup tab contains preferences associated with the startup of the software startup Add Samples Analysis Users Project ie Open Blank Project t Open Previous Project Element Description Open Blank Project option button Sets the software so that upon launch a blank Project window opens Open Previous Project option button Sets the software so that upon launch the last Project viewed will open GeneMapper ID Software v3 1 User Guide Appendix C Project Window Software Interface Add Samples Tab The Add Samples tab contains preferences associated with the adding of samples Startup Add Samples Analysis Users When
148. ced Mode The Advanced mode provides the user with the same analysis parameters available in GeneScan Software v3 7 1 Windows NT OS Basic Mode The Basic mode is not recommended for human identification applications because it allows only for analysis using limited parameters consisting of a user defined minimum peak height threshold GeneMapper ID Software v3 1 User Guide l 3 Appendix GeneMapper ID Software Verification AmpFZSTR Kits Samples Instrumentation Hardware and Software Used for Verification Testing AmpF STR Kits The following AmpF STR PCR Amplification kits were used to Used amplify the samples used in the verification testing e 4 Dye Kits COfiler Green I Profiler Profiler Plus SGM Plus e 5 Dye Kits Identifiler SEfiler Sample Types Table I 1 lists the DNA sample type the total number of samples Used amplified and the total number of alleles available for evaluation and comparison for the verification of GeneMapper ID software Table l 1 Samples used for software verification DNA Sample Type NumberctSamples Number of Alleles Single source 16 7422 Mixture 4 3678 Degraded 3 2402 Inhibited 3 1318 Low quantity 3 21 12 High quantity 3 2 22 l 4 GeneMapper ID Software v3 1 User Guide AmpF amp TR Kits Samples Instrumentation Hardware and Software Used for Verification Testing Thirty two human genomic DNA sample
149. cel Figure 5 23 General Tab of the Plot Settings Editor GeneMapper ID Software v3 1 User Guide 5 43 Chapter 5 Using GeneMapper Manager Sample Header The Sample Header tab controls what is viewed in the header above Tab each sample electropherogram in the Samples Plot view Plot Settings Editor Column Sample File Sample Name Sizing Quality os Oe ia 3 5 Figure 5 24 Sample Header Tab of the Plot Settings Editor The Show button displays all selected rows as visible checked and the Hide button displays all selected rows as hidden unchecked 5 44 GeneMapper ID Software v3 1 User Guide Section 5 4 Plot Settings Tab Genotype Header The Genotype Header tab controls what is viewed in the header above Tab each Genotype electropherogram in the Genotypes Plot view Plot Settings Editor Offscale Sharp Peak M One Shc Allele M e einge Peaxaact a a m snitPeaka 10 outorBinalele a B fpeakHeimran Figure 5 25 Genotype Header Tab of the Plot Settings Editor The Show button displays all selected rows as visible checked and the Hide button displays all selected rows as hidden unchecked GeneMapper ID Software v3 1 User Guide 5 45 Chapter 5 Using GeneMapper Manager Sizing Table Tab The Sizing Table tab controls which columns are displayed in the 5 46 Plot Settings Editor column OvelSample Peak Sample File Mame Marker Allele Size Height Area Data Point
150. certain actions or Add and Remove Rows as desired Edit Add Mew Row Ctrl A4 Remove Row Ctrl R 4 Click OK to save and close the Size Standard Editor or click Cancel to close without saving 5 71 Chapter 5 Using GeneMapper Manager 5 72 GeneMapper ID Software v3 1 User Guide Options Dialog Box This chapter covers IMM OGUCHOM eacee eean ne rie ete aS Aes ao TA 6 2 HID Analysis Options 0 0 0 0 0 eens 6 3 Siac Nab Aint def Sa eas Stent ob a ei So ates ieee 6 4 Add Samples Tab y n0025 445528 Oo APE RDO AGAR DES Oa ass 6 5 Analysis MAD tae Hehehe oe eae a ae ea eee eae ees 6 7 WS6IS AD c0 acid ere aa ak eee eee ane eee konere E 6 9 GeneMapper ID Software v3 1 User Guide 6 1 Chapter 6 Options Dialog Box Introduction The Options dialog box contains several tabs for setting preferences associated with various software features You can use the tabs in the Option dialog box to e Set project preferences e Set default analysis parameter preferences e Set default post analysis display preferences e Create New Users and Passwords Note Each registered user has his or her own set of preferences When you set these options it affects only the user currently logged in 6 2 GeneMapper ID Software v3 1 User Guide HID Analysis Options HID Analysis Options Overview In preparing GeneMapper JD software to analyze data from AmpF STR kits view and set options for e Startup of GeneMapper ID so
151. ch return and removal of the software from buyer s systems terminate the license to use the software and refund the buyer s purchase price for the software If there is a defect in the media covered by the above warranty and the media is returned to Applied Biosystems within the ninety 90 day warranty period GeneMapper ID Software v3 1 User Guide Limited Product Warranty Applied Biosystems will replace the defective media Applied Biosystems does not warrant that the software will meet buyer s requirements or conform exactly to its documentation or that operation of the software will be uninterrupted or error free Warranty Period Any applicable warranty period under these sections begins on the Effective Date earlier of the date of installation or ninety 90 days from the date of shipment for software installed by Applied Biosystems personnel For all software installed by the buyer or anyone other than Applied Biosystems the applicable warranty period begins the date the software is delivered to the buyer Warranty Claims Warranty claims must be made within the applicable warranty period Warranty The above warranties do not apply to defects resulting from misuse Exceptions neglect or accident including without limitation operation outside of the environmental or use specifications or not in conformance with the instructions for the instrument system software or accessories improper or inadequate maintenance by the
152. crosoft Excel In addition you can not change the order in which the columns are listed What is displayed in the genotypes table is determined by the table setting selected in the Table Setting drop down box in the project window The table settings are created in the GeneMapper Manager refer to Chapter 5 Section 5 3 You can simply switch back and forth between table settings by changing the setting in the drop down box Keep in mind that the table setting change affects both the samples table and the genotypes table Note The default settings are designed to display two alleles per marker Therefore if you are working with mixtures and need to display more than two alleles per marker you will need to change the Number of Alleles within the Genotypes Tab of the Table Setting Editor GeneMapper ID Software v3 1 User Guide 7 27 Chapter 7 Analyzing and Interpreting Data Sorting a You can sort the table by any present column header Genotype Table To sort a Genotype table 1 Ifthe genotypes table is not already displayed click the genotypes tab in the main project window or go to View gt Genotypes 2 Select Edit gt Sort The following window will appear You can sort up to three criteria ir Ascending C Descending G Ascending S Descending G Ascending Sample File Z C Descending OK Cancel 3 Select the criteria to sort by from the drop down boxes and choose ascending A Z 1 2 3 or de
153. ct this user name on launching GeneMapper ID software the name is used to identify the preferences set by you Note User names must be unique If the new user name is not unique an alert message will be displayed Note Users added here are for this GeneMapper ID software installation Users are visible to all other users The names are converted to lower case and passwords are not case sensitive GeneMapper ID Software v3 1 User Guide Users Tab Element Description Change Opens the Change Password dialog box and allows Password you to change the password for the selected user button F Change Password Change password for user qm Password Confirm Password GeneMapper ID Software v3 1 User Guide 6 11 Chapter 6 Options Dialog Box 6 12 GeneMapper ID Software v3 1 User Guide Analyzing and Interpreting Data This chapter covers IMIHOCUCHOM seess cenres ne a Sakis wd S a AMO ny bcd nes 7 2 Considerations for HID Analysis 00 000 eee 7 2 Analyzing Samples Files 0 00006 cc eee eee eee ene 7 3 Interpreting Sample PQVs Sizing Quality and Precision 7 11 Size Match EdiOr t0 t es eoeneeea hes ectes cg EE 7 14 Se Galline CURVE sareren EEEE a pebraree na onee ean 7 19 Viewing Sample Data 0 0 0 0 eee eee 7 23 Genotypes lable 44742 ierre nen A AE eos eet e a eee 7 27 Interpreting Genotype PQVs and Genotype Quality 7 30 Concordance
154. culated ratio at these loci are just above GeneScan software v3 7 1 or just below GeneMapper ID software Advanced mode the defined stutter ratio Two examples are given below Macro Panel Calculated Stutter Setting Ratio Sample Type After Analysis LL LL 5 c ons ey e 9 m iz T X me 14 14 ai ai Stutter Calculated Stutter Defined in Macro Panel Setting 3100 1to9 D18851 13 422 14 2472 17 07 Identifiler mixture GeneMapper ID 13 4182 14 2466 16 95 3100 Genotyper THO1 6 72 7 1161 620 6 Profiler Plus GeneMapper D 6 674 7 1157 579 6 a The alleles and peak heights shown in italics were filtered out in GeneMapper JD Observation 5 0 01 2 20 364 alleles of the alleles from offscale data analyzed with GeneScan software v3 7 1 and Genotyper software v3 7 Windows NT OS did not filter the actual stutter because of the limitations of the Genotyper software Filtering compares Peak A to the next labeled peak Peak B The label is removed from Peak A the stutter peak if Peak B the true allele meets two criteria GeneMapper ID Software v3 1 User Guide Results of Verification Testing Peak B is higher than peak A by the specified percentage and Peak B is within the specified proximity size in base pairs range relative to peak A These two observations for verification did not meet these criteria due to an abundance of A product resul
155. d Genotypes View menu items and descriptions continued Item Description Off scale Peak Indicator Controls whether or not offscale peaks are highlighted in an electropherogram Checked when active Note When highlighting of offscale peaks is enabled a magenta bar overlays any offscale peak in an electropherogram as shown in the figure below An offscale peak is a peak whose height exceeds the dynamic range of the collection instrument Samples Plot File Edit View Tools Alleles Help Plot Setting HID Genotyping X fy Panes fi SiS e BEE LAMM 44 Vi E AES E Sample Name Pana Qo SQ Izle E TE X 182 61 _Y 4821 Bin 18 Marker AA Dyes Controls the dye colors shown in the Plot window Checked when active Multiple dye colors may be selected Note This feature is not available when viewing microsatellite genotypes in the Genotype Plot window A v Blue Dye Ctri 1 Plots how Green Dye Ctrl 2 Tables ko ov Yellow Dye Ctri 3 Table Filter ho wRedDye Ctri 4 w Header s Orange Dye Ctrl 5 w hlarker Range Ctri R w All Dyes Ctrl 6 GeneMapper ID Software v3 1 User Guide Plot Window Menus View menu items and descriptions continued Item Description Plots Controls the way electropherograms of samples are shown in the Sample Plot window Combine Dyes Ctrl Tables Separate Dyes Ctrl 8 Table
156. d In File Edit Analysis View Tools Help FA BS cee co Sy M L Hl Faj GD table setting HiD Table E E Project info Raw Data EPT Data E E Casework CO_Ladder fsa H CO_Control fsa OO Ladder fsa 0 1000 2000 3000 4000 5000 6000 CO_Sample1 tsa CO_Sample2 fsa PP_Cortrol fsa PP_Ladder fsa PP_Sample1 fsa PP_Sample2 fsa data point 1088 rfu 4883 Progress Status J stropa Plot scaling features in the Raw Data view e Zoom in on one area by placing the cursor in either the X or Y axis label areas and click and drag The software zooms in on the area of selection e Return to a full scale plot by double clicking in the label area again The plot window is restored to full scale e Scale the Y axis to maximum Y or a user defined value by selecting Y Axis Scale from the View menu GeneMapper ID Software v3 1 User Guide 3 7 Chapter 3 Using the Project Window Use the Raw Data view to evaluate the following e baseline problems or noise could result in poor size calling e start and stop points for analysis EPT Tab Clicking the EPT electrophoresis power and temperature tab in the Samples view when the Sample file is selected displays a window that shows the voltage power current and run temperature associated with that sample F GeneMapper ID 3 1 Casework Project gmid Is Logged In File Edit Analysis View Tools Help
157. d fragments that may run anomalously 11 P27 11 Best Fit 2nd Order Curve AO 9 7716235E 01 Al 4 1 220838E 01 AZ 4 856055E 06 Size Calling Curve Cubic Spline Interpolation I I I I I I 1500 2000 2500 S000 3500 4000 Data Point Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper ID software v3 1 Possible Local Sizing Inaccuracy Mobility of any DNA fragment can be affected by its sequence and by secondary and tertiary structure formation If any internal size standard fragment has anomalous mobility the Cubic Spline method may exhibit local sizing inaccuracy For example Assume that a standard fragment is close in molecular length to an unknown sample fragment Assume further that the standard fragment runs anomalously The Cubic Spline method assigns the official value to this standard fragment even though it may be slightly incorrect The size of the unknown fragment is then likely to be calculated incorrectly as well Note This method does not determine the amount of sizing accuracy error GeneMapper ID Software v3 1 User Guide B 31 Appendix B Software Genotyping Algorithms Local Southern The Local Southern method determines the sizes of fragments by Method _ using the reciprocal relationship between fragment length and mobility as described by E M Southern 1979 Best Fit 2nd Order Curve AO 1 990206E 02 Al 2
158. d when the table contains data Analysis Menu The commands in this menu control analysis and display of the data in a Project and access to a number of other windows that support the Project window such as the Analysis Manager and Panel Manager The following table is provided as a reference for the Analysis menu Display Flots wiew Tools Help Ctri L Size Match Editor Analyze Analyze Selected Samples Analyze All Analyze Markers Analysis Method Editor size Standard Editor Bring Errors to Top Cirl R Cirl Shitt Fe Cirl Bring Mon concordant Samples to Top Item Description Enabling Display Plot Ctrl L Opens the Plot window Enabled when one or more samples or records are selected Size Match Editor Opens the Size Match Editor window Enabled only when sized samples are present in the Project GeneMapper ID Software v3 1 User Guide Appendix C Project Window Software Interface Item Description Enabling Analyze Analyses all samples eligible Enabled when Ctrl R for analysis The symbol is Samples tab is visible shown in the Status column and at least one sample in the table requires analysis Analyze Analyzes selected samples Enabled when Selected only Samples tab is Samples visible Analyze All Analyzes all samples Enabled when Ctrl Shift R whether or not the Status Samples tab is column reports t
159. dated Genotyper software calls these off ladder alleles or OL Allele and allows the end user to edit the alleles accordingly This example was chosen specifically for the off ladder variant for verification purposes This sample was used for both single source and mixture amplifications for all kits therefore it contributes to 18 observations of the 12 2 allele being called Male 23_2ng Profiler_Phus_vl a 30 100 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 Figure l 6 GeneMapper ID software genotyped D18851 locus with the 12 2 allele ES Genotyper 3 7 Plots PROFILER_PLUS_ 1 gta i E la HD Fie Edit Analysis Category Table Views Macro Window Help lal Ix bse 20 40 60 30 100 360 380 400 420 440 120 140 160 180 200 220 O 240 260 289 300 320 340 0851179 D21511 Z Ns FS Male_23_2ng fsa 3 Green htale_23_2ng 30 2 16 Figure l 7 Genotyper software genotyped D18S51 locus with the OL allele GeneMapper ID Software v3 1 User Guide l 15 Appendix Platform and Ampf str Kits GeneMapper ID Software Verification Observation 4 0 04 8 20 364 alleles of the alleles from the mixture data used in comparing GeneScan software v 3 7 1 to GeneMapper D software Advanced mode produced eight alleles where the minor contributor s alleles that are 4 bp smaller than the major contributor s alleles i e in the stutter position are filtered since the cal
160. denoted by red circles allele caller outputs are denoted by green black and blue asterisks Note that consensus between multiple callers virtually assures that the calls are correct In certain cases some algorithms have not made a call since they have determined that the data is too complex to act on as in 1 and p Here the blue asterisks show the calls transmitted to you Low quality values are reported because in both cases the first algorithm did not call and in 1 the black caller is not in agreement with the blue However despite these conditions the calls are correct The low quality values alert you to potential problems such as the spurious peak in 1 and the high background in p a 1251659_27 b 51597_24 c 85258_24 d 125 5 1066 1354 h 7S486_21 1118 488 1386 W k 1283107 1011 1926 AIA i g AEN A m 1271_14 o 118987_3 1426 B 4 GeneMapper ID Software v3 1 User Guide GeneMapper ID Software Genotyping Algorithms Quality Value The Quality Value Determination algorithm is used to create Process Determination Quality Values PQVs PQVs are reported by data analysis and are Algorithm an aid to finding and fixing problems in sample preparation and analysis These values are the end results reported by the PQV system The quality value determination algorithm equation is the following MQ x AQ GQ MQ x POV x PQV x xPQVy e PQVs are weighted fro
161. differences in relative fluorescent units RFU for single source DNA samples and low quantity DNA samples for a total of 3483 alleles Table l 4 Peak detection evaluation AmpF STR aa gear E A of PCR Instrument Mode Number Ee ee FA Amplification Platform Used of Alleles _ Height Height Peak Kit Difference Difference Height RFU RFU Difference Profiler Plus 310 Genetic Classic 610 0 15 0 13 0 3 and COfiler Analyzer Macintosh Identifiler Operating 551 0 8 0 26 0 4 System Profiler Plus 310 Genetic Advanced 610 0 16 5 71 2 6 and COfiler Analyzer NT Operating Identifiler System 551 0 20 4 61 1 1 Profiler Plus 3100 Genetic 610 0 11 3 69 0 6 and COfiler Analyzer Identifiler 551 0 16 3 34 1 1 The GeneMapper ID Classic mode was designed to produce similar results to GeneScan Software v 3 1 2 Macintosh OS so a laboratory could maintain its current analysis parameter settings The average peak height difference measured is less than 1 RFU for the Classic mode a very insignificant difference Although a maximum difference of 15 RFU was observed in one allele the maximum percent difference for this allele is only 0 3 The maximum peak height difference measured in all samples evaluated is 20 RFU observed in a single source DNA sample using the Advanced mode l 8 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Peak Height Comparison Example Dat
162. displayed In Figure I 10 one allele from each locus was Removed during the functionality test Further D8S1179 allele 12 was selected and its Allele History is displayed In the Allele History table GeneMapper ID software v3 1 Created this allele In performing this functionality test for verification the Allele History table states that this allele has been Deleted GeneMapper ID Software v3 1 User Guide Results of Verification Testing NY t File Edit Yiew Tools Alleles Help Plot Setting HID Genotyping PAM Paes z ME MT a FEN al SA E i ef hae W dP Wdentifiler_v1 L 2 lle gl lt 175 72 Y 149 F allele History J Allele Marne User Mame Modification Date Action Comments 39 66 fa986 12 ni 2003 11 06 11 35 30 0 Crested GeneMapper HID Allele Calling Algorithrn i 39 66 1 2 ni 2003 1 07 10 46 22 14 Deleted Funetionalty_Test Export OK Cancel Figure l 10 Deleted Allele and the Allele History table Locus The fourth parameter evaluated in the Peak Detection and Concordance Genotyping category is the locus concordance between duplicate Evaluation amplifications of the same sample and locus concordance between the same sample using different AmpF STR kits The requirement is that samples must have the same name Samples that have the same name are evaluated if the non concordance feature is selected 254 GeneMapper ID Softwa
163. drive s and removable media drives 2 In Files tab navigate to folder containing sample files Select the folder then click the Add to List button Folder is added to the Samples To Add field Individual samples within a folder can be selected and added Selecting the folder adds all samples 3 Click the Add button Add Samples to Project window closes and the project window becomes visible Samples are added to the bottom of the Project window table Edit Menu The commands in the Edit menu of the Add Samples window are used to manage the contents of the window The commands are described in the table below ig via SEER All they Add Samples to List Ctrl D Eles WElETE Clear All b Options C 38 GeneMapper ID Software v3 1 User Guide Add Samples to Project Dialog Item Description Enabling Select All Ctrl A Selects all the items of a given type Enabled when a single valid selection has been made e g sample file Add Samples to List Ctrl D Clear Delete Adds the selected item s from the Files GeneMapper D database Clears deletes the selected item s in the Samples to Add list Enabled when a sample file or folder with files in the Files GeneMapper D database is selected Enabled when a valid selection is made Clear All Hierarchical menu Clears all sample files and folders from the specified field e Samples to add e Criteria Note
164. ds Tab F size Standard Editor k a x Template File Info File CO Control fsa Run Date amp Time 0002 11 30 00 00 00 0 Dye Red Analysis Method HID_ classic Size Standard Description Harme JROX_Size_Standard Size Standard Table Size Standard Data Points 1 33291 2 3623 100 0 a 3954 139 0 4 4074 150 0 5 4157 160 0 G a511 200 0 OK Cancel Figure 5 32 The Size Standard Editor in Classic Mode The Size Standard Editor in Classic mode shows an electropherogram and a table of peaks for the dye color and sample selected The software assigns a number to each peak found in the electropherogram in order from left to right You should be able to recognize the peak pattern of the standard in the electropherogram Note You can only change the peak size value in the right column of the table You cannot change or rearrange the peak numbers Note If too many peaks display in the electropherogram or the baseline is too high you might need to adjust the analysis parameters 2 Enter in a name and description for the size standard GeneMapper ID Software v3 1 User Guide 5 69 Chapter 5 Using GeneMapper Manager 5 70 3 Specify the peaks of the standards and their sizes Click the peak you want to define either in the electropherogram or in the table or click on a row and the corresponding peak is highlighted Note For easier viewing you can zoom in on the electrophe
165. e database To display the Table Settings tab select Tools gt GeneMapper Manager gt Table Settings GeneMapper Manager Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Name Last Saved Owner Description HI Table 2003 06 12 14 38 22 grid Mew Oper Save S Inport Export Delete Figure 5 16 Table Settings Tab of the GeneMapper Manager GeneMapper ID Software v3 1 User Guide 5 27 Chapter 5 Using GeneMapper Manager Table Settings Tab Elements Default Table Settings 5 28 Table 5 7 Description of elements on the Table Settings Tab Item Description Name column Table Setting name Last Saved column Owner column Date Time stamp displaying when the object was last saved User name Description Description of the object column New button Opens the Table Setting Editor window Always enabled Open button Opens the editor for selected table settings Enabled when a single object is selected Save As button Displays the Save As dialog box Enabled when a single object is selected Import button Displays a dialog box for Importing objects Always enabled xml file Export button Displays a dialog box for Exporting selected objects Enabled when one or more objects are selected xml file Delete button Deletes the selected object s Done button Closes the GeneMapper
166. e Gl B Bf 100 HA Fe gama ee eee 1 version 2 Chernistry Kit 3 BinSet Name 4 Panel Name 5 Marker Mame E ASR A 8 Marker Mame 9 ASR 10G 11A 12 Marker Mame T3 ASR 14G 15 Marker Mame 16 ASR MF IT 18 Marker Mame 19 ASR 0c EET 22 Marker Mame 23 ASR ff ar M44 Hi Sheeti Sheet 4 GM 3 1 oNP dermo oNPdemo bplexdemo Omer 246 246 Zomer of 2r 20 5 Samer 30 4 308 4 dAmer 46 5 46 6 52 Mer 53 1 53 1 at BO mer 61 4 61 4 26 1 26 1 Greer 29 9 24 1 Blue 24 4 Green 402 40 2 Blue 45 0 45 5 Red 552 S46 Yellow 55 2 Red BS Bo Yellow ie ee ee eee 1 2 3 4 Column Description 1 Bin name 2 Bin size range minimum 3 Bin size range maximum 4 Bin color GeneMapper ID Software v3 1 User Guide Formats of Panel and Bin Text Files Import and The Panel Manager imports and exports two text file data formats Export Files panel data and bin set data These files are tab delimited and the following rules apply e Files to be imported must have the correct number of columns in the correct order e On import panel and bin set names are checked for uniqueness in the GeneMapper ID database and markers are checked for uniqueness within the panel that will contain them An alert is presented if a name is not unique and such a file cannot be imported e If any errors are found durin
167. e Info Ctrl F 4 Raw Data Ctrl F 2 EPT Data Ctrl F 3 Full view Ctrl AAXIS SC ale Labels Bins Ctrl Shitt B Peak Positions Allele Changes Controls To Top Off scale Peak Indicator Dyes Plots Tables Table Filter Header Marker Range Ctrl R Marker Indicator Cirl k r F F F Marker hargin View menu items and descriptions Item Description Toolbar Controls whether or not the Plot toolbar is displayed at the top of the Plot window This item is checked when active Sample Info Provides quick access to the Sample Info data tab for the Ctrl F1 selected electropherogram See Chapter 3 Using the Project Window Raw Data Provides quick access to the Raw Data tab for the Ctrl F2 selected electropherogram See Chapter 3 Using the Project Window GeneMapper ID Software v3 1 User Guide 8 7 Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued Item Description EPT Data Ctrl F3 Provides quick access to the EPT Data tab for the selected electropherogram See Chapter 3 Using the Project Window Full View Ctrl Automatically resets the X and Y axes of the electropherogram s currently viewed to their full scale size File Edit View Tools Alleles Help Plot Setting HD Genotyping IMM rsi ABA RTF ER a Ee Fi 24
168. e Plot Window 00c cc cece eeeeeee 8 2 Plot Window Toolbars 0 00 cece ee eee ee eee 8 4 Plot Window Menus c0 cece ene eeeueeueeee 8 5 GeneMapper ID Software v3 1 User Guide 8 1 Chapter 8 Using Plot Windows Samples and Genotypes About the Plot Window 8 2 Introduction Purpose of the Plot Window Displaying Plot Windows The Samples and Genotypes plot windows allow you to visually assess your data If genotype calls are created you can view and edit allele calls assigned by the GeneMapper ID software algorithms Samples Plot Window The Samples plot window allows you to e View electropherograms on a per sample basis e View a Sizing or genotypes table for sample s shown e Overlay all samples to determine size standard quality e Edit markers and bins e View the relationship between controls and samples e Edit allele calls Genotypes Plot Window The Genotypes plot window allows you to e View electropherograms on a per genotype basis e View all genotype calls for a given marker e Edit markers and bins e View the relationship between controls and samples e Edit allele calls Display the plot window from the GeneMapper Project window To open the plot window 1 Highlight any number of samples or genotypes from the project window by selecting the row number or sample file name 2 Click m Display Plots on the toolbar or Analysis gt Display Plots When you se
169. e beginning of the run and in a flat part of the baseline and record the data point values for both the start and stop points in the flat part of the baseline of the data point range Note When choosing the start point do not include primer peaks in the data point range refer to Eliminating Primer Peaks on page 5 59 Enter these values in the next step when generating the new matrix refer to page 5 60 2 Return to the Project window by selecting the run folder in the navigation pane Eliminating Both the primer peaks and the data peaks are displayed when you Primer Peaks view the raw data of your matrix standards Any time you run dye labeled samples on a gel ABI PRISM 310 Genetic Analyzer or capillary ABI PRISM 377 DNA Sequencer you have excess dye labeled primer in the reaction The primer peak displays as the first peak usually off scale because it is in molar excess Eliminate the primer peak when making a matrix by choosing the start point after the primer peak in a flat area with a stable baseline Note To create a good matrix you need at least five fragments in each color GeneMapper ID Software v3 1 User Guide 5 59 Chapter 5 Using GeneMapper Manager Generating a New Matrix Creating a To create a GeneMapper ID software matrix GeneMapper I D 1 Select GeneMapper Manager gt Tools Then click the Software Matrix Matrices tab The Matrices page opens GeneM apper Manager 5 60 Gen
170. e files p PP_Control tsa l oe PP Ladder fsa PP_Samplet fsa i PP_Sample2 fsa GeneMapper ID Software v3 1 User Guide Navigating in the Project Window Once you have added files to the Project window If you Then select the project item in the Navigation pane all samples in the project are displayed in the table select a run folder in the Navigation pane select a sample file in the Navigation pane samples in the selected Run folder are displayed in the table you have access to sample information raw data and EPT data Click the tab corresponding to the type of data you want displayed in the right pane GeneMapper ID Software v3 1 User Guide 3 13 Chapter 3 Using the Project Window Viewing the The following table lists how to display information in a Project Project Window window Contents To hide the Open the Clear the And GeneMapper ID software displays the Navigation pane View menu Show Project window without the navigation pane from the Project Navigator window check mark _ GeneMapper ID 3 1 Casework Project gmid ls Logged In Alternative File Edit Analysis View Tools Help Ctrl Shift N ek oo al A LLL Ss l Samples Genotypes Status Sample Mame Sample Type Specimen Cater Analys 1 i OL Control Sample no export AD_ Ch J 4 CO Ladder Allelic Ladder no ex Th HID _CI k Samplez Samp
171. e no export F Advanced cotier v cE F_HID_ Gd To ho eo Ladder er Allelic Ladder er no export F Advanced COfier_ v CE FH F H Gq ENE Samplet Sample na exp export oto F_Adh Advanced eea a O OO 1 i gt Progress Status SYS step With the Project node highlighted in the left navigation pane all sample files will be displayed To display only the sample files associated with one analyst highlight the first or second folder The first folder is associated with the lead analyst and the second folder is associated with the second read You can duplicate run folders on the server to give these folders different names to provide more obvious differentiation Note To import the same sample files the project must be saved closed then reopened If you do try to import the same sample files without closing it first an error message displays and the files are not imported K 2 GeneMapper ID Software v3 1 User Guide 7 Select the folder of samples just imported then analyze and edit the data and save the project Both sets of analysis are saved within the same project 8 To compare results Select the Project node in the left navigation pane of the main window Select Genotypes tab gt Analysis Menu gt Non concordant sample to the top Any non concordant calls then display at the top GeneMapper ID v3 1 Casework gmid Is Logged In Fie Edit Analysis View Tools Help e s
172. e sort is ascending or and descending in alphabetical numerical order Descending buttons OK button Closes the Sort dialog box and performs the sort Cancel Closes the Sort dialog box Does not perform the sort button C 46 GeneMapper ID Software v3 1 User Guide GeneScan Size Standards This appendix covers ATPOCUCH ORS sara nesenie aa 6 akin who elated a ead i D 2 GeneScan 120 Size Standard 000 000 0000 ee eee D 3 GeneScan 350 Size Standard 00000000 D 4 GeneScan 400HD Size Standard 0 0 00005 D 5 GeneScan 500 and GeneScan 500 250 Size Standards D 7 GeneMapper ID Software v3 1 User Guide D 1 Appendix D GeneScan Size Standards Introduction The GeneMapper ID Software Version 3 1 comes with several ready to use GeneScan size standard definition files that you can choose from to analyze fragments run on ABI PRISM genetic analyzers The following table lists the ready to use size standards stored in the GeneMapper D database If using size standard Select this GeneScan 120 LIZ GS120LIZ GeneScan 400HD GS400HD GeneScan 500 GS500 GeneScan 500 250 GS500 250 GeneScan 500 LIZ GS500LIZ GeneScan 500 250 LIZ GS500 250 LIZ Note The GeneScan 350 size standard is not automatically stored in the database however it is provided in the GeneMapper gt Size Standards folder for import D
173. e v3 1 User Guide Interpreting Genotype PQVs and Genotype Quality Override _ Invalidate Genotype Quality Genotype Quality This feature allows you to quickly reset the Genotype Quality to 1 0 and all PQV icons will be greyed out Since the allele call has not changed the AE allele edit box will not be checked To use the Override Genotype Quality feature in the sample plot 1 Show the Genotype table 2 Select the Genotype Quality cell of a genotype 3 Right click the selected cell to open the Override Genotype Quality dialog box 4 Click Yes The Genotype Quality now passes with a value set to 1 0 and the PQV icons are greyed out in the Genotypes table To use the Override Genotype Quality feature in the genotype plot 1 Select the sample header of a plot 2 Right click the header to open the Override Genotype Quality dialog box 3 Click Yes The Genotype Quality now passes with a value set to 1 0 and the PQV icons are greyed out in the Genotypes table Genotype Quality The Genotype PQVs are used to calculate the overall Genotype GQ Quality GQ The user can set level of importance of PQVs in the overall calculation of the Genotype Quality GQ in the Quality Flags Tab of the Analysis Method Editor refer to Chapter 5 Note The GQ for a specifc marker in a ladder file represents the genotype quality for that marker in that ladder file One of the components involved in GQ calculation is the ladder matching
174. e v3 1 User Guide Section 5 6 Overview Section 5 6 Size Standards Tab Size Standards Tab The Size Standards tab in the GeneMapper Manager window contains a table of size standards stored in the database To display the Size Standards tab select Tools gt GeneMapper Manager gt Size Standards Figure 5 30 displays the four default size standards that are loaded with the GeneMapper ID software on an analysis computer only and Figure 5 31 displays the standards load when GeneMapper ID software is loaded on a computer running 3100 3100 Avant Data Collection software v2 0 F GeneMapper Manager ee Meade Tre ee Metrces Sename Description Se a See Se a7 F HD_essoo 2003 10 14 19 40 5 gmid Basic Advanced Factory Provided CE_65_HD_esso0 2003 10 14 19 40 5 gmid BasiciAdvanced Factory Provided CE_F_HID_ SS00 2003 10 14 19 40 5 gria BasiciAdvanced Factory Provided Mew Oper Save Aa Import Export Figure 5 30 Standards for a stand alone analysis computer GeneMapper Manager Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Mame Name Lat Saved Owner Type Description GSSDDLIZ 2003 10 20 14 32 72 arrid Pa Factory Provided GSS00t 250 LIZ 2003 10 20 14 32 72 gtd BasiciAdvanced Factory Provided Mew Oper Save AS Import Export Figure 5 31 Standards for 3100 3100 Avant system computer using Data Collec
175. eMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix 2 Click the New button on the Matrices tab The Matrix Editor dialog box opens E j Matrix Editor 3 Enter in a Matrix name and description if desired 4 Choose the number of dyes from the Number of Dyes drop down list If 5 dyes are selected a button for O data is added to the bottom of the list GeneMapper ID Software v3 1 User Guide 5 61 Chapter 5 Using GeneMapper Manager The B G Y R and O buttons represent dye colors Choose the file associated with the appropriate dye color a Click a button to display a pop up menu b Use the pop up menu to access a sample file to link to each of the dye labeled primers c Choose the sample file that represents the dye color for that button Enter the start point that you determined when choosing a data point range in the Start At field Refer to Choosing a Data Point Range on page 5 59 Enter the total number of data points to include in order to calculate the matrix in the Points field In most cases leave the default value unless you must exclude a portion of your data Note You must have at least five peaks to make a matrix Click Create to generate a new matrix Click OK to save and close the Matrix Editor dialog box Matrix Example The following is an example of the matrix results showing the values used to calculate the overlap correction
176. eMapper JD software after the trained analyst completed editing Peak Detection Concordance 0 09 0 04 0 01 o Observation 1 E Observation 2 o Observation 3 E Observation 4 a Observation 5 Figure l 5 Peak detection concordance observations e Observation 1 99 18 20 197 20 364 alleles of the alleles compared are the same whether analyzed with GeneScan and Genotyper software or GeneMapper ID Software v3 1 Note The remaining 0 82 of alleles are described in Observations 2 through 5 e Observation 2 0 68 139 20 364 alleles of the alleles from low quantity degraded and inhibited DNA samples fell below the defined 50 RFU peak amplititude threshold setting in GeneMapper ID software Advanced mode All of these alleles were analyzed to l 14 GeneMapper ID Software v3 1 User Guide Results of Verification Testing be just below 50 RFU resulting in the peak not being called in GeneMapper D software whereas they were detected as 50 RFU or slightly greater in GeneScan software As previously described the GeneMapper JD software Advanced mode has an overall slightly lower peak height 2 6 or less compared to GeneScan software v3 7 1 e Observation 3 0 09 18 20 364 alleles of the alleles from one single source DNA sample at the D18S51 locus with the 12 2 allele are called in GeneMapper JD software since the panels have been updated with known variants The macros for Genotyper software have not been up
177. eak detection Description of the Analysis Method Editor Process Project window GeneMapper Manager window algorithm for your application In the Peak Quality tab set minimum requirements In the Quality Flags tab adjust PQV and threshold settings Click OK to save Figure 5 7 Analysis Method Editor Process Analysis Method Editor window 5 12 GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab Accessing the Access the Analysis Method Editor in the following ways Analysis Tae e Select Tools gt GeneMapper Manager gt Analysis Methods Tab gt New or Open e In the Project Window click the Samples tab Click New Analysis Method to create a new method or double click an existing method to edit it e Select a sample in the project window and click the Analysis Method Editor icon on the toolbar Edit the currently selected samples analysis method Note The Analysis Method Editor icon 1s not active unless a sample with an analysis method is selected Analysis Method Editor HID x Peak Detection Algorithm advanced ag Ranges Feak Detection Analysis Sizing Partial Range Partial SIZES start Pt 2700 Start Size 75 stop Pt reno stop Size 450 Smoothing and Bazelining Peak Amplitude Thresholds B fo mfo G o ofo no EL wh hin Peak Halt width 50 50 k Polynomial Degree 3 f 5 0 0 0 0 Smoothing C plone E Light t Heavy
178. ection Polynomial Degree and Peak Window Size Parameters 2 000 cece cece eee eee B 17 Optimizing Peak Detection Sensitivity Example 1 B 20 Optimizing Peak Detection Sensitivity Example 2 B 22 Optimizing Peak Detection Sensitivity Example 3 B 24 Peak Detection Slope Threshold for Peak Start and Slope Threshold for Peak End Parameters 0005 B 25 Slope Threshold Examples 0 0000 e eee eee eee B 27 Size Calling Methods for Classic and Advanced Modes B 29 Appendix C Project Window Software Interface Overview of the Project Window 0 0 c eee eee eee eee C 2 Project Window Menus 000 eee eee ee ees C 10 Project Window File Menu Dialog Boxes 00000 0 ee C 27 Printing Tables and Electropherograms 000000 ee eae C 34 Add Samples to Project Dialog 0 00 eee eee eee C 37 Project Window Edit Menu Dialog Boxes 0005 C 45 Appendix D GeneScan Size Standards FI EPOCIIC HOI ser tos cz See aire orate estan sel ea ee eee eae eae Gon sen a ae eee D 2 GeneScan 120 Size Standard 0 ce D 3 GeneScan 350 Size Standard ce ee D 4 GeneScan 400HD Size Standard 0 ce ee D 5 GeneScan 500 and GeneScan 500 250 Size Standards D 7 Appendix E Converting Macintosh Sample Files Converting Macintosh Sample Files 0 00000 e eee eeee E 2 vi GeneMapp
179. ects tab Analysis Methods tab to create custom analysis methods for use in analyzing sample files Table Settings tab to create new profiles hide and show table columns and filter the table entries rows in Project windows Plot Settings tab to create a custom plot profile for viewing data Matrices tab to create a matrix file Size Standards tab to create a size standard e Chapter 6 Options Dialog Box describes how to set up default analysis settings customize analysis settings and create new users and passwords e Chapter 7 Analyzing and Interpreting Data describes how to Set HID analysis options for analysis interpreting sample PQVs sizing quality and precision Use the Size Match Editor to examine size standard electropherograms edit the identification of size standard peaks and view size calling curves GeneMapper ID Software v3 1 User Guide Organization of the User Guide View Genotypes table interpreting genotype PQVs and genotype quality Perform concordance checks e Chapter 8 Using Plot Windows Samples and Genotypes describes how describes how to examine and interpret electropherograms within the samples and genotypes plot windows e Chapter 9 Exporting Table Data describes how to use the export and CODIS export features e Chapter 10 Printing Data describes how to print data Appendices The following appendices contain reference information e Appendi
180. ediate value again in a window whose width Beta is set using the Baselining options and centered at each data point i GeneMapper JD software baselines an electropherogram by subtracting the baseline from the raw electropherogram Troubleshooting the Baseline Window The following table describes what happens if the baseline window is either too small or too large Using Causes a small baseline window size the baseline to creep into the peaks resulting in shorter peaks in the analyzed data the baseline to ride too low resulting in elevated and possibly not baseline resolved peaks a large baseline window size Size Calling GeneMapper JD software uses these methods to determine the Method Options molecular length of an unknown fragment Description of the size calling method parameter options Item Description 2nd Order Least Squares and 3rd Order Least Squares Both Least Squares Methods use regression analysis to build a best fit sizecalling curve Cubic Spline Interpolation Forces the sizing curve through all the known points of the selected size standard Local Southern Method Determines the sizes of fragments by using the reciprocal relationship between fragment length and mobility Global Southern Method Similar to the Least Squares Method in that it compensates for standard fragments that may run anomalously GeneMapper ID Software v3
181. eight Data Point items in the label box the GeneMapper D and Size labels software v3 1 AE Comment Drop down menu Shows the last allele item in the label box edit comment Note If you delete an allele the AE Comment is not displayed in the genotype table is visible in the Allele History The AE Comment is only displayed for changed alleles Show data type Check box Displays label prefixes prefixes such as al for allele Call Show type of edit Check box Displays type of edit made to the allele If an allele is manually edited the label will display changed or deleted Invert mutant labels Check box Identifies mutant bin If a bin is labeled as a mutant bin in the Panel Manager then the label color is inverted to distinguish it from normal bins 5 48 GeneMapper ID Software v3 1 User Guide Section 5 4 Plot Settings Tab Table 5 13 Labels tab menu items continued Name Type Description Label Color Drop down menu Controls the color of the labels From the drop down menu select one of the following Black and White Dye Color All text and border in color Dye Color Border text in black border in color GeneMapper ID Software v3 1 User Guide 5 49 Chapter 5 Using GeneMapper Manager Display Settings The Display Settings tab controls how each plot setting will Tab automatically open when a customer chooses t
182. el Locus The New Kit Panel Locus command changes its name and function depending on the item selected in the Panel Manager navigation pane as follows e New Kit This menu item is enabled when the Panel Manager root node is selected in the navigation pane and allows creation of a new kit folder When creating a new kit 1 Enter in a unique kit name 2 Select the appropriate kit type microsatellite or SNP from the drop down menu 3 Click OK to add kit to navigation pane GeneMapper ID Software v3 1 User Guide 4 7 Chapter 4 Using Panel Manager e New Panel This menu item is enabled when a kit folder is selected in the navigation pane Use New Panel to create a new panel folder When creating a new panel 1 Select the New Panel name in the right hand window table and enter in the desired name Names must be unique within the database 2 Repeat to add multiple panels to a kit 3 Press Enter then click Apply after entering a Panel name to update the navigator pane e New Marker This menu item is enabled when a panel folder is selected in the navigation pane Use New Marker to create a new locus When creating new loci 1 Select the New Marker row in the right hand window table and enter in the appropriate information for each column Refer to Marker Table View on page 4 36 for more information 2 Repeat to add multiple markers to a panel 3 Press Enter then click Apply
183. elected for the matrix Description A description of the matrix New button Opens the Matrix Editor dialog box Always enabled Open button Opens the editor for a selected matrix Enabled when a single matrix is selected Save As button Displays the Save As dialog box Enabled when a single matrix is selected GeneMapper ID Software v3 1 User Guide 5 53 Chapter 5 Using GeneMapper Manager Table 5 14 Description of elements in the Matrices tab continued ltem Description Import button Displays a dialog box for Importing matrices Always enabled Note The extension must be mtx Export button Displays a dialog box for Exporting the selected matrix Enabled when one or more matrices are selected Note The extension must be mtx Delete button Deletes the selected matrix s Always enabled Done button Closes the GeneMapper Manager Always enabled About Matrices Introduction There are two dye labeling chemistries currently available to prepare nucleic acid samples to use the GeneMapper JD software on ABI PRISM instruments 5 end labeling Dye Sets F or G5 Each chemistry has a set of dye labels that fluoresce at different wavelengths when excited by a laser The wavelengths are During data collection on the separated ABI PRISM 310 Genetic Analyzer by a spectrograph into a Known ABI PRISM 377 DNA Sequencer spectral pattern across a detection 377XL or
184. em Description Marker Indicator Ctrl K Controls whether or not the red triangles at the bottom of the electropherogram which indicate the marker allele size range are displayed Marker Margin Opens the Marker Margin dialog box to allow you to determine how many base pairs beyond the allele size range are displayed in the electropherogram Note This option is only enabled when viewing the Genotype plots The value can be pre configured through the Plot Setting Editor F Marker Margin x Enter marker margin n basepairs a GeneMapper ID Software v3 1 User Guide 8 21 Chapter 8 Using Plot Windows Samples and Genotypes Tools Menu The Tools menu provides access to the GeneMapper Manager and the Plot Settings Editor To ols Genehappertanager Ctri M Plot Settings Ctrl T Tools menu items and descriptions Item Description GeneMapper Opens the GeneMapper Manager to allow creation or Manager editing of various project settings Used to create a new Plot Ctrl M Setting profile See Chapter 2 Getting Started with GeneMapper ID Software for more information F GeneMapper Manager l x Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Project Last Saved Owner of Samples 2003 11 13 grid 3 CaseVYork_2 CaseWork_3 2003 11 13 grid Rename Save As Import Export
185. en select Edit gt Delete from Project Sizing Quality Note Different peak detection algorithms give different Sizing Quality SQ results Therefore refer to the appropriate section below for instructions on interpreting SQ Classic Peak Detection Algorithm To examine the size standard 1 Select Edit gt Select All and click Size Match Editor to view the size standard for all samples 2 Notice that the Sizing Quality value equals 0 5 GeneMapper ID Software v3 1 User Guide 7 11 Chapter 7 Analyzing and Interpreting Data m Sizing Quality Size Matches Size Calling Curve Sizing Quality 0 5 Override SQ 439 150160 3500 3700 3900 4100 4300 4500 4700 4900 5100 5300 5500 5700 5900 6100 3 View the peak assignments for each sample a Press the down arrow key to scroll through the samples on the left side of the Size Match Editor screen b Confirm that the size for the peaks in the size standard are correctly assigned for each sample 4 Select Edit gt Override All SQ to override the SQ value for all samples Notice that the Sizing Quality is changed to lt 1 0 gt which indicates that the size standard was verified by the user ii Quality Size Matches Size Calling Curve Sizing Quality lt 1 0 gt 429 150160 3500 3700 3900 4100 4300 4500 4700 4900 3100 5300 5500 5700 5900 6100 7 12 GeneMapper ID Software v3 1 User Guide Interpreting Sample PQVs
186. en you add samples the panel for all samples is set to the value in the text box When the option button is selected the following choices may be made e Leave the text alone which selects the existing name or e Click the field to present the Select a Panel window to select an existing panel The Select a Panel window contains the list of panels from the GeneMapper D database Ea Select a Panel El AmpFLSTR_Panels_v1 pr Blue sne Green pee Profiler mna Profiler Plus_v1 odie COfiler_v1 oi SGM Plus w1 When you add samples the panel for all samples is set to the value in the Sample Sheet comment column Set Sample Type to e Sample type for all samples option button and drop down menu e Read from Data collection Info field option button When you add samples the Sample Type for all samples is set to the value chosen from the Sample Type drop down menu contains choices for Sample Control Allelic Ladder and Primer Focus When you add samples the GeneMapper D software does a best guess match comparing the contents of the field specified in the Sample type box with the list of sample types Sample Control Allelic Ladder Negative Control and Primer Focus e Ifa match is found then the Sample Type property for the sample is set to that sample type e fa match is not found then the Sample Type property for the sample is set to
187. equally and the curve 1s not constrained to go through any specific point The software can analyze a large range of fragment sizes with this method DNA fragments that are Are sized using not bracketed within the size a second order least squares curve standard curve extrapolation bracketed within the size standard the method that was chosen curve For best results use a standard that brackets all the fragments of interest GeneMapper ID Software v3 1 User Guide B 35 Appendix B Software Genotyping Algorithms B 36 GeneMapper ID Software v3 1 User Guide Project Window Software Interface This appendix gives detailed descriptions of the features of the GeneMapper ID Software Version 3 1 application including details of each major application window This appendix covers Overview of the Project Window 000 cece eee C 2 Project Window Menus 3 6 0 3426dh0 k08 host oboe debe C 10 Project Window File Menu Dialog Boxes C 27 Printing Tables and Electropherograms C 34 Add Samples to Project Dialog 00000 08 C 37 Project Window Edit Menu Dialog Boxes C 45 GeneMapper ID Software v3 1 User Guide C 1 Appendix C Project Window Software Interface Overview of the Project Window Menus and The Project window has a number of menus and dialog boxes Dialog Boxes associated with it The drop down menus available
188. er ID Software v3 1 User Guide Appendix F Appendix G Appendix H Appendix Appendix J Appendix K Glossary Index Transferring Data and Backing Up Data IWanSieMming Dale a vaienliGa steno iaden eles ERG Dade wade webs F 2 Backing Up Data and Cleaning the Database F sample Sheets and Plate Records Information from the Data Collection Sample Sheet or Plate Record G 2 Troubleshooting Sheet Troubleshooting Sheets 22i i2 ivie ei Fitna dees dde wee ewido H 2 GeneMapper D Software Verification Summary of GeneMapper ID Software Verification 2 AmpF STR Kits Samples Instrumentation Hardware and Software Used for Verification Testing 20 0 c eee eee eee eee 4 Results of Verification Testing 2 00 ees l 7 GONCIUSION 264 ocwewaddowse 00 64 a aa A l 39 References ununnan nananana Sete a feeds Mae Gade art a es l 40 Software Warranty Computer Configurations 00 arie ea eas Swe eee as J 2 Limited Product Warranty 0 0 0 eee ees J 2 Workflow for Storing Sample Files Projects and Second Reads GeneMapper ID Software v3 1 User Guide vii viii GeneMapper ID Software v3 1 User Guide Preface How to Use This Guide Purpose of This The GeneMapper ID Software Version 3 1 User Guide provides Guide reference information about the GeneMapper JD software for general use and customization For specific step by step instruc
189. er wi El Protiler Plus v4 4 Profiler_Plus_v1 none Fl COfiler w 5 COfier none H E SsGM Plus vt 6 SGM _Plus v1 none A f idertifiler v1 T Identifiler_v1 none se tiler_sv1 Protiler_Plus_Cot ra SEfiler_v1 none Fl cotiler COD v g Profiler_Plus_CODIS_v1 none H uere cons E 10 COfiler_CODIS v1 none 4 a Identifiler_CODIS v none OK Cancel Apply Conceptually the hierarchy of panel data is as follows e Kit chemistry kit or product e Panel e Locus e Bin For AmpF STR data analysis different bin sets are not needed for different instrument platforms The panels and bins supplied by Applied Biosystems for AmpF STR data analysis are optimized to accommodate all instruments that support AmpF STR data analysis The GeneMapper ID software requires that kits panels and bin sets are unique throughout the database and that locus names are unique within a panel The Panel Manager enforces these requirements during user entry editing of names and data import Since the Panel Manager allows users to add comments as needed no history or audit trail of panel data changes is kept Such comments provide a way for users to track changes and the reasons for the changes Each kit panel and locus has a comment field GeneMapper ID Software v3 1 User Guide Panel Manager Panel Manager Overview The toolbar icons enable specific Panel Manager actions similar to Toolbar their associ
190. erlap e This feature serves as a warning for an allele calling error e This is used for HID only NB Narrow Bin e A flag Pass Il or Check A is automatically displayed when a peak is present at a position within 0 5 basepairs from a bin and no peak is present inside that bin e The benefit is to capture peaks that fall outside of bin boundaries due to experimental variations e This flag is used for SNP only GeneMapper ID Software v3 1 User Guide Using Process Component Based Quality Values The PQV flags on the Genotypes page continued Column Description DP Double Peak e A flag Pass Ii or Check A is automatically displayed when two peaks of the same color in the same bin have a ratio 20 5 minor peak height major peak height e This flag is used for SNP only e Not editable GQ Genotype Quality e A red octagon E is shown when the analysis for a sample is of low quality e Pass f Check or Low Quality flags are set by the Analysis Method Not editable Below is a representation of sample peaks with the allele quality AQ assigned AQ AQ See the following page for information about the mathematical formula used to calculate the Genotype Quality from a sample represented by the image above GeneMapper ID Software v3 1 User Guide Appendix A Process Quality Values The PQV flags on the Genotypes page continued Column Descr
191. ersion programs to prepare Macintosh computer generated fragment analysis sample files for transfer to a Microsoft Windows based format and vice versa This appendix covers Converting Macintosh Sample Files 0 E 2 GeneMapper ID Software v3 1 User Guide F 7 Appendix E Converting Macintosh Sample Files Converting Macintosh Sample Files About Converting E 2 Sample Files Installing Conversion Programs Applied Biosystems created two conversion programs that prepare sample files for transfer from a Macintosh computer to computers running Microsoft Windows NT operating systems and vice versa These sample file conversion programs run only on a Macintosh computer The sample file conversion programs do not perform the file transfer from computer to computer They set attributes of the files so that they can be used on the destination computer For example when transferring a fragment analysis sample file from a Macintosh computer to a computer running the Windows operating system a file extension 1s required and the conversion program adds fsa to the sample file name For more detailed information on how these conversion programs function refer to the SimpleText file entitled About Conversion Programs located in the same folder as the sample file conversion programs To install the sample file conversion programs on a Macintosh computer 1 Insert the GeneMapper ID software CD ROM int
192. es E TA O E E A E E tab Mame Last Saved Owner Type af G5 HID GSs500 2003 08 06 13 10 O gril BasiciAdvanced 377 F_HID_ 3500 2003 08 06 13 10 O gria BasiciAdvanced CE_G5_HD_GS500 2003 05 06 13 10 0 grid BasiciAdvanced CE_F_HD_GSs00 2003 08 06 13 10 0 gin BasiciAdvanced HID_Classic_GSS00 250 2003 08 09 15 58 2 gmnid Classic Hew REN Seve ME Import Expor Size Match Analysis gt Size Size Match Editor Editor Match Editor Alternative Size Match Editor Click the icon a a x Eg Size Matches size Calling Curve LU Sizing Quality lt 1 0 gt Gyende Sa OK Cancel AppIy 3 16 GeneMapper ID Software v3 1 User Guide Navigating in the Project Window To see the Select And GeneMapper ID software displays the Table Setting Editor Tools gt Table Setting Editor Alternative Press Ctrl T If samples have been imported you can click the icon Note To create a new table select Tools gt GeneMapper Manager Then select the Table Settings tab and click New Table Setting Editor Table Setting Editor General Samples Genotypes Samples Table Settings Column Settings Font Settings Show Column Filtering Content 1 m Status Show All Records N A a 20r Er Show All Records i 3 p7 Sample Name Show All Records 4 7 Sample ID Show All Records 5 pm Comme
193. ess Quality Values A 8 The PQV flags on the Genotypes page continued Column Description SPA Single Peak Artifact e A flag Pass Ii or Check A is automatically displayed when the marker signal contains single peaks due to some problem in electrophoresis e The flag is triggered when there are no peaks present within a two base pair range prior to an allele peak e This feature detects the absence of stutter peaks which indicates nonmicrosatellite peaks e This flag is used only for microsatellite dinucleotide markers only e Not editable Bin Out of bin allele e A flag Pass Ii or Check Ad is automatically displayed when the called alleles peak apex is out of the bin boundary e Triggering this flag reduces the final PQV Genotype value The default is 20 0 2 multiplier of what it would be otherwise e For HID this is labeled OL off ladder alleles e This flag is used for microsatellite analysis e Not editable PHR Peak Height Ratio e A flag Pass oor Check A is automatically displayed when there are two alleles present and the ratio between the lower allele height and the higher allele height is below a certain level e fthere are more than two alleles present the calculation iterates through all the peak pairs e PHR can be set in the Peak Quality tab of the analysis method Analysis Manager The default is 50 e For microsatellite markers the ratio is calcul
194. estions FAQs Submit a question directly to Technical Support Order Applied Biosystems user documents MSDSs certificates of analysis and other related documents Download PDF documents Obtain information about customer training Download software updates and patches In addition the Support page provides access to worldwide telephone and fax numbers to contact Applied Biosystems Technical Support and Sales facilities GeneMapper ID Software v3 1 User Guide xi Preface xii GeneMapper ID Software v3 1 User Guide About GeneMapper ID Software This chapter provides an overview of the GeneMapper ID Software Version 3 1 software features supported chemistry kits and this user guide This chapter covers Overview of GeneMapper ID Software 00040 1 2 Steps Required to Use GeneMapper ID Software 1 5 Organization of the User Guide 000 0c cee eee 1 6 GeneMapper ID Software v3 1 User Guide 1 1 Chapter 1 About GeneMapper ID Software Overview of GeneMapper D Software Supported Instruments 1 2 GeneMapper D Software Version 3 1 is an automated genotyping software solution for forensic paternity and database data analysis and other genotyping needs The GeneMapper ID software combines functions of the GeneScan Analysis Software and the Genotyper Software and it includes additional features and enhancements The GeneMapper ID Software Version 3 1 i
195. ete A warning alert is shown Click OK to remove the setting The Samples Table Settings column labeled Filtering shown in the figure below is used to filter or set the type of record or display presented for a Project window column Table Setting Editor General gt amples Genotypes Samples Table Settings Column Settings Font Settings Font arial x Show Column 1 9 Fittering status Show All Recorda MA Sample File Show All Records a a m Sample Name Show All Records 4 m Sample ID Show All Records E m Comments Show All Records G W Sample Type Show All Records T al Specimen Category Show All Records 5 r Analysis Method Show Al Records g m Panel Show All Records ee Show Hide OK cancel Figure 5 21 Filtering Column of the Table Settings Editor GeneMapper ID Software v3 1 User Guide 5 35 Chapter 5 Using GeneMapper Manager Samples View Column Filtering Settings Note When you apply these settings to filter parameters be aware that when no instance of the type specified exists the Sample Table displays blank Return to the Table Manager and reset the filter The following information describes how to use the filter settings for the Samples Column Settings The default Show All Records can be changed to one of the following Table 5 10 Samples view filter settings Column Name Filter Settings Status Sample File Analyzed o
196. eters Work How to Use These Parameters Use the Polynomial Degree and the Peak Window Size settings to adjust the sensitivity of the peak detection You can adjust these parameters to detect a single base pair difference while minimizing the detection of shoulder effects or noise Sensitivity increases with larger polynomial degree values and smaller window size values Conversely sensitivity decreases with smaller polynomial degree values and larger window size values The peak window size functions with the polynomial degree to set the sensitivity of peak detection The peak detector computes the first derivative of a polynomial curve fitted to the data within a window that is centered on each data point in the analysis range Using curves with larger polynomial degree values allows the curve to more closely approximate the signal and therefore the peak detector captures more peak structure in the electropherogram The peak window size sets the width in data points of the window to which the polynomial curve is fitted to data Higher peak window size values smooth out the polynomial curve which limits the structure being detected Smaller window size values allow a curve to better fit the underlying data Use the table below to adjust the sensitivity of detection To Polynomial Window Size Degree Value Value Increase sensitivity use Higher Lower Decrease sensitivity use Lower Higher Gene
197. ethod of analyzing size standards and performing peak detection Analysis Method Editor HID Ei General Allele Peak Detector Peak Quality Quality Flags Peak Detection Algorithr classic Ranges Feak Detection Peak Amplitude Thresholds Bio Ro G 50 50 Y 50 Analysis Sizing Fun Range jal Sizes Stant Hi fo Start Tal Stop Pi i gg Stop Sise f g Data Processing V Baseline m MutiComponent Smoothing 7 None i Light c Heavy Min Peak Half Width 3 pts Split Peak Correction Ino Peak Correction T Correction Limit 30 pts Size Calling Method 2nd Order Least Squares 3rd Order Least Squares Cubic Spline Interpolation Local Southern Method Global Southern Method Factory Defaults OK Cancel Figure 5 13 Peak Detector Tab of the Analysis Method Editor Classic Peak Detection Algorithm GeneMapper ID Software v3 1 User Guide 5 21 Chapter 5 Using GeneMapper Manager Peak Quality Tab 5 22 The ranges and peak amplitude thresholds are identical to those for the advanced algorithm Refer to Advanced Peak Detection Algorithm on page 5 19 for details Refer to Appendix B Software Genotyping Algorithms for additional information Note Applied Biosystems recommends that you use the default settings for all other analysis parameters for AmpF STR data analysis The Peak Quality tab allows the user to set thresholds that
198. ethods Always enabled Export button xml file type Displays a dialog box for Exporting selected analysis methods Enabled when one or more analysis methods are selected Delete button Deletes the selected analysis method s Done button Closes the GeneMapper Manager GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab Default Analysis GeneMapper I D software v3 1 contains the following standard Methods Applied Biosystems profiles Analysis Bin Set Detection pelaut Type Algorithm Microsatellite Microsatellite AmpF STR_Bins_v1 Classic Advanced SNP SNP None Basic Genotyping Genotyping Note These profiles can be altered but should not be deleted They are intended to be used as models for customizing new analysis methods Note The GeneMapper ID Software v3 1 Human Identification Analysis Tutorial PN 4335523 shows you how to create methods that are used as a guide for AmpF STR data analysis GeneMapper ID Software v3 1 User Guide 5 11 Chapter 5 Using GeneMapper Manager Analysis Method Editor Introduction The Analysis Method Editor allows you to create or edit analysis methods to be used for analyzing your sample files The analysis method determines the type of application being analyzed and allows for customization of the peak detection algorithm and quality value settings ae In the Peak Detector tab choose a p
199. etting Started with GeneMapper ID Software Note Refer to Appendix A for PQV column information GeneMapper ID Software v3 1 User Guide C 9 Appendix C Project Window Software Interface Project Window Menus Introduction Command Activation and Special Terms File Menu The tables in this section describe the following Project Window menus Topic See Page File Menu C 10 Edit Menu C 12 Analysis Menu C 13 View Menu C 15 Tools Menu C 17 Help Menu C 26 The commands in the Project Window menu can be activated in two ways e Clicking the command in the menu or e Using the special key combinations shown to the right of the name in the menu The commands are described in the table below gem Edit Analysis View Tools Mew Project Open Project reve ni EL Save ProjectAs Add samples to Project Export Table Print Log Out Exit Cirl h Ctro iils Cirl l ctrl E Ctrl F Ali F 4 GeneMapper ID Software v3 1 User Guide Project Window Menus Item Description Enabling New Project Clears sample files results and Always enabled Ctrl N project name from the Project except when a Open Project Ctrl O window displaying a blank Project If previous project has pending changes the following alert message is displayed Do you want to save changes Yes No Cancel Displays the Open Project dialog box
200. ew User dialog box ra CALI User re Mew User name Password Confirm Password To add a user name for GeneMapper ID software click New User to open a dialog box and then a Type a user name into the dialog box b Enter a password using only alphabetic characters c Confirm the password d Click OK to assign the user name and close the dialog When you select this user name on launching GeneMapper ID software the name is used to identify the preferences set by you Note User names must be unique If the new user name is not unique an alert message will be displayed Note Users added here are for this GeneMapper ID software installation and is visible to all other users The names are converted to lower case and passwords are not case sensitive Opens the Change Password dialog box and allows you to change the password for the selected user F Change Password Change password for user gm Password Confirm Password GeneMapper ID Software v3 1 User Guide C 25 Appendix C Project Window Software Interface Help Menu The Help menu provides access to online help and to the About GeneMapper JD software window GeneMapper Help F1 About Genehapper 49 Item Description Enabling GeneMapper Help Opens the GeneMapper ID User Always F1 Manual PDF file using Acrobat enabled Reader Does not open the tutorial guides About Opens the About GeneMapper ID
201. ewing the data in different plot configurations e g five samples showing all dye colors in five panes using three modes of zooming no zoom medium zoom and tight zoom ranged from 0 15 seconds to 0 37 seconds The printing test was only conducted on one printer different printers may have different results Data Handling The fourth category presented is data handling Test Parameters The tested parameters are described below 1 Import and export functionality test of projects analysis methods table settings plot settings matrices and size standards 2 Verify CODIS export functionality using both export extension files CMF 1 0 and CMF 3 0 Import Export The first parameter tested in the data handling category is the import Functionality and export functionality of projects analysis methods table settings plot settings matrices and size standards Projects with their associated settings 1 e analysis methods table settings plot settings matrices if applicable and size standards should be exported and archived regularly in order to maintain the database Further the associated settings must accompany the exported project in order to accurately assess the original analyses Note When the project is exported the sample files are not linked Results The ability to export and import projects analysis methods table settings plot settings matrices and size standards is 100 functional GeneMapper ID So
202. fa z ER A e B all Mesije E E aE E B D8 1179 DISI D75820 CSFIPO 100 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 D3S1358 THOI DI35317 0 220 2 DI6ess39 D2 13 30 240 250 260 270 280 290 300 310 320 330 340 350 100 110 120 130 140 150 160 170 180 130 200 21 800 400 hs fe b b Sample Identifier vi C D19543 FFA POX 18551 100 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 A nA Re A SL WR ER ee eR O E A FGA 100 110 120 130 4140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 lze aj Note Bins are enabled in Separate Dyes mode only for Microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot kit samples When you turn off a specific dye color for SNaPshot kit samples the bins for that color turn off also GeneMapper ID Software v3 1 User Guide Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued ltem Description continued Samples Plot File Edit View Tools Alleles Help Bins Combine Dyes Mode O x rA E ho Sample Name Pand Plat Setting HD Genotyping rones 1 NNT a ET al EE SQO SQ Ideniifile_vl 1179 CRIE chi Nomen D8 CSFI
203. fferent Copying the matrices will and Analysis ensure that the correct matrix name is stored in the sample file The proper matrix is required for accurate analysis of ABI PRISM 310 Genetic Analyzer and the ABI PRISM 377 DNA Sequencer sample files To transfer the matrix from one computer to another l Click the Matrices tab in the GeneMapper Manager window 2 Select Matrix to Export 3 Click Export 4 5 Select a name and location for the matrix Transfer the mtx file to the data collection computer GeneMapper ID Software v3 1 User Guide 5 63 Chapter 5 Using GeneMapper Manager Where to Store Store exported matrix files mtx intended for use by Data Collection Exported Matrix software in Files D AppliedBio Shared Analysis Sizecaller Matrix If Data Collection and Analysis are installed on different computers the location is the same Remember to copy the matrix from the analysis computer to the Data Collection computer Note GeneMapper D software matrices are stored in the Oracle database Causes for Bad Matrices lf an Error There are two possible causes for the error messages shown in the Message following table Displays If Take this action you designated the wrong files reassign the matrices Refer to Assigning the Matrix to Samples on page 5 63 the signal is too weak to make a rerun the matrix standards matrix 5 64 GeneMapper ID Softwar
204. file raw data from each of the matrix standard files so you can decide where to choose the start point for the data point range To view raw data l 2 3 Open a new GeneMapper ID Project Import Samples by selecting File gt Add Samples to Project Select the files you want to view a Navigate to the disk directory containing the samples for creating a matrix b Select the files you wish to use c Click Add to List Click Add to import the files into a new project and close the Add Samples to Project window The Project window reopens with the imported files displayed in the Samples tab Select a sample file Select Raw Data from the File menu GeneMapper ID Software v3 1 User Guide Section 5 5 Creating and Evaluating a Matrix What to Look For In the raw data display of the sample files verify the following in the Raw Data Display e Data peaks are present in all four or five of the matrix standards e There are no anomalies e The baseline is stable flat between peaks e Peaks are on scale peaks do not exceed 8191 relative fluorescent units and the peaks of the dye of interest have a value of at least 200 If peak data does not show these characteristics refer to Causes for Bad Matrices on page 5 64 for possible interpretations of your peak data Choosing a Data To choose a data point range Point Range 1 Move the cursor well away from the primer peak in a region at th
205. formation When bins are imported into a kit GeneMapper ID software checks for bins that fall outside of their marker allele size ranges If bins overlap or fall out of the marker range the Import Bins Options dialog box will open Import Bins Options a xj ated select Option C Increase marker range to accomodate all bins Delete all overlapping and out of bounds bins OK Cancel Use the Import Bins Options dialog box to choose how GeneMapper ID software handles the bins being imported The numbered lines in the picture above refer to the individual numbered descriptions in the following figures GeneMapper ID Software v3 1 User Guide 4 9 Chapter 4 Using Panel Manager 1 The Adjust marker range amp delete out of bounds bins option shown below adjusts the marker range to accommodate any bins that overlap the marker boundaries and will delete all bins that fall completely outside the marker size range 1 Adjust marker range and delete out of bounds bins iil Marker Range 98 114 A B c D E F G Marker Range 100 110 2 The Increase marker range to accommodate all bins option shown below increases the marker allele size range to accommodate all overlapping and out of bounds bins 2 Increase marker range to accommodate all bins i jiji Marker Range 88 116 A B c D E F Marker Range 100 110 4 10 GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands 3
206. ftware e Analysis settings when adding samples to a project e Analysis displays e Users of GeneMapper ID software Viewing and Note These options are active only for the user currently logged into Setting Options the software To view and set options 1 Select Tools gt Options to open the Options dialog box 2 From the Startup tab view the default startup options Project Open Blank Project C Open Previous Project Note Later you may select Open Previous Project to open the last project you analyzed using GeneMapper JD software GeneMapper ID Software v3 1 User Guide 6 3 Chapter 6 Options Dialog Box Startup Tab The Startup tab contains preferences associated with the startup of the software startup Add Samples Analysis Users Project ie Open Blank Project Open Previous Project Element Description Open Blank Project Sets the software so that upon launch a blank option button Project window opens Open Previous Project Sets the software so that upon launch the last option button Project viewed will open 6 4 GeneMapper ID Software v3 1 User Guide Add Samples Tab Add Samples Tab The Add Samples tab contains preferences associated with the adding of samples Startup Add Samples Analysis Users When adding new samples automatically Set Analysis Method to gt Ho Advanced for all samples Read from the Sample
207. ftware v3 1 User Guide l 37 Appendix GeneMapper ID Software Verification I 38 CODIS Export Functionality The second parameter tested in the data handling category is the CODIS export functionality test One advantage of GeneMapper JD is the ability to add extension files that are recognized by the Combined DNA Index System CODIS software These samples files can be exported and uploaded into the CODIS software Ten sample files both single source and mixture samples using the AmpFISTR Identifiler AmpFISTR Profiler Plus and AmpFISTR COfiler PCR Amplification Kits were successfully imported using both CODIS v 5 6 and CODIS v 5 7 The FGA locus in the Identifiler PCR kit has nine 9 extended alleles Applied Biosystems 2001 When performing the verification of GeneMapper JD one 1 extended allele at the FGA locus was not recognized The CODIS Change Control Board has been informed of this finding GeneMapper ID Software v3 1 User Guide Conclusion Conclusion Applied Biosystems performed verification of GeneMapper ID v3 1 to assess the performance robustness and feature designs of four categories 1 Peak detection and genotyping 2 Algorithms 3 Workflow and 4 Data Handling Within these categories over 20 000 alleles and software features were verified for 24 specific test parameters The data for this verification were evaluated and are summarized in this appendix The verification was performed usi
208. g Printing Sizing Plots Similar to Genescan Using the Predefined AmpF STR Sizing Plot Setting Below is a representation of how the data will print when using this plot setting Sample Name Pand 500 So CO Conve Cone vl x i H B 1 CO Control fsa 115 42 105 O03 S770 ontrol fsa gE CO _Control t 0351356 15 123 52 i _Cortrol fsa B 4 Co Control t 249 44 65 431 4337 r B 5 ga Control fsa Dt 65535 j1 253 35 1 457 i 1598 4970 r 5 5 COL Control fsa Dt 65539 2 257 34 1 arr i 2672 5003 m le coL Control fsa 02 48 182 932 3647 m ez 2 col Control fsa AMEL x 03 4 2383 i 5110 S656 E bB 2 COL Control fsa 3st 355 4 19 5 201 a 1 2 62 3808 1 953 12450 3845 CO _Control ftsa THO 179 16 1077 BESS 4328 To print this view l Within the samples tab of the main project window select the samples in the table you want to display in the printouts Select Analysis gt Display Plots Select AmpF STR Sizing from the Plot Setting drop down list Refer to Chapter 5 for information on importing plot settings into the GeneMapper Manager Select File gt Print leaving all page set up options as default GeneMapper ID Software v3 1 User Guide 10 7 Chapter 10 Printing Data Printing Genotype Plots Similar to Genotyper Using the Predefined AmpF STR Genotyping Setting The allelic ladder and control samples as defined in the sample type will be printed first if multiple samples a
209. g 14 5 Sample4 Sample no export HID_Advanced identifier _v1 LIZ_size_standard Databasing 4 6 Samples Sample no export HID_Advanced dentifiter_v1 LIZ_size_standard Databasing 14 llc s gt l Analysis Completed Sto Note You can hide and show columns in the table using Table Setting GeneMapper ID Software v3 1 User Guide C 3 Appendix C Project Window Software Interface Samples View Columns The columns of the Samples view are explained in the table below F GeneMapper ID 3 1 Database gmid Is Logged In File Edit Analysis view Tools Help e S BPS ww L l U E p Table setina ameFLSTR Table m8 8 Project Samples Genotypes Databasing Status Sample Name Sample Type Specimen Cater Analysis Method Panel Size Standard Matrix Run Name iin 1 ID_Control Positive Control no export Identifiler Identifiler_v1 LIZ_size_standard Databasing A 2 ID_Ladder Allelic Ladder no export HID_Advanced Identifiler_ v1 LIZ_size_standard Databasing A 3 ID_Neg_Critrl Negative Contre no export HID_Advanced Identifiler_ v1 LIZ_size_standard Databasing A 4 Sampe3 Sampe Convicted Offer HID_Advanced identifier v1 LIZ_size_standard Databasing a 5 Sample4 Sample no export HID_Advanced Identifiler_ v1 LIZ_size_standard Databasing A 6 Samples Sample no export HID_Adyvanced Identifiler_ 1 LIZ_size_standard Databasing A
210. g import the Panel Manager does not accept data from the file e Lines beginning with are comments and are ignored on import e Both Panel and Bin set file formats have a header line with a GeneMapper I D software version string Version GM JD v 3 1 If the first line of a file to be imported does not begin with Version GM then GeneMapper JD software handles the import as a GeneMapper JD software 1 x panel import If the file is in GeneMapper D software 1 x format then it is imported without errors If the file is in GeneMapper ID software 2 x or 3 x format the file will not be imported and the following error alert will be presented The format of this file is wrong Correct the file and try again If the data type label does not match the type specified in the import command the file is not imported and an alert is presented A PanellmportLog txt file is generated during import of a panel or bin set and will list any errors encountered This file is found in the GMdistribution app folder on your hard drive GeneMapper ID Software v3 1 User Guide 4 33 Chapter 4 Using Panel Manager Panel Table View Introduction Selecting the Panel Manager icon or text in the navigation pane left pane shown below shows a table of the current Panel Manager folders or kits in the right portion of the window Selecting a particular kit produces a list of panels in the right portion of the window This list is the Panel Ta
211. gs These settings contain the valid columns for either Samples or Genotypes data e Show column List with an editable check box for each Project column controlling whether the column is shown or hidden e Column column List of column headings for the Project window e Filtering column List of filtering properties for each Project window column e Content column Sets what information is displayed in each Project window column as selected by the appropriate filtering method N A indicates that no comments can be made for these rows Filtering The filtering properties for a Project table column Properties are controlled in the Filtering and Content columns Summary Modifying these settings updates the property in the Column Filter table For more information see For more specific Filtering Controls on page 5 35 information see Filtering Controls on page 5 35 OK Closes the Table Setting Editor window and applies any pending changes to the Project window Contents of the Table Setting Editor are saved to the GeneMapper D database Cancel Closes the Table Setting Editor window without making any pending changes Show Selects all highlighted rows as visible Hide Selects all highlighted rows as hidden GeneMapper ID Software v3 1 User Guide Section 5 3 Table Settings Tab Allele Settings The Allele Settings box on the Genotypes page of the Table Setting Ed
212. hat setting Plot Settings Editor General Sample Header Genotype Header Sizing Table Labels Display Settings When Gpening The Plot indo Use the display settings last used for this plot Use these display settings For both Sample and Genotype plots Panes 1 Peli ia fi el pels AN S Basepairs Axis Scale individually M Toolbar M Show Off scale Le Bee M For Genotype plot only hlarker Margin 5 bp Cancel Figure 5 28 Display Settings Tab of the Plot Settings Editor Description Show hide plot header Show hide marker range Show hide marker indicators Show hide bins Hide labels Horizontal labels EEEBEE 5 50 GeneMapper ID Software v3 1 User Guide Section 5 4 Plot Settings Tab Description Vertical labels Show hide peak positions Bring Don t bring controls to top Show hide allele changes PE EE Show Hide Individual Dyes or All Dyes No table rrr kkk k k k Sizing Table Genotype table Combine Dyes Separate Dyes Overlay All FEEE E You can select Use the display settings last used for this plot button to have the Samples and Genotypes plot views display with your previous settings or select Use these display settings to adjust the display with the current settings on this page These icons are the same as the ones in the
213. hat the visible sample is eligible for analysis including those previously analyzed The following alert message is displayed Analyze all samples This may change your existing results This action cannot be undone OK Cancel Analyze Analyzes only the marker s Enabled when Markers you select within each Samples tab is sample Analyze Markers visible Dialog Box on page C 15 Analysis Allows you to change the Enabled when a Method Editor settings for analysis sample with an analysis method is selected Size Standard Editor Allows you to edit base pair and dye settings Enabled when one or more samples or records are selected Low Quality To Top Ctrl B Sorts all Low Quality SQ or GQ column samples or results records to the top of the table Enabled when analyzed samples are present in the table Non concordan t Samples to Top Brings non concordant samples to the top of the analysis page GeneMapper ID Software v3 1 User Guide Project Window Menus Analyze Markers Dialog Box The Analyze Markers dialog box enables you to select which marker or set of markers separated by commas will be analyzed within each sample Fl Analyze Markers Wai 2 This analysis option is designed for Microsatellite and SMaPshot analyses Enter marker names separated by commas cane This option is only enabled when you are in the Project window Samp
214. he Minus A cut off value The Minus A peak is displayed as a shoulder peak to the parent peak 1 base pair less than the main allele Three different values were chosen to test 2772 alleles in three high quantity DNA samples and 16 alleles in one single source sample for Minus A cut off at ratios of 0 20 and 30 The results Figure I 16 on page I 26 show that the algorithms implemented in GeneMapper JD software properly filtered the percentages entered in the analysis method editor GeneMapper ID Software v3 1 User Guide l 25 Appendix GeneMapper ID Software Verification Genotypes Plot File Edit View Tools Alleles Help Plot Setting HID_Genotype z E Panes fa zir i ha ESS AL Sample Name Pand Marker e are no controls in this selection rr Profiler Pius vi 351358 A A E E SI OS BIN PHR LPH SPU AN BD CE OVL GQ I 0 Minus A Cut Off offscale 38_ ng Profile_Plus_v1 B3sisse Q A A m go A m go k 20 Minus A 7 Cut Off i 30 Minus A Cut Off Figure l 16 Example of a high quantity 8ng DNA sample tested using three Minus A cut off values Minus A is filtered with increasing cut off values I 26 GeneMapper ID Software v3 1 User Guide Results of Verification Testing PQVs Parameters 3 to 7 in the Algorithm Testing category are the individual PQVs for the off scale flag verification the off
215. he Find dialog box Tooltip Find Ctrl F Q a Opens the Print dialog box Tooltip Print Ctrl P GeneMapper ID Software v3 1 User Guide 3 19 Chapter 3 Using the Project Window 3 20 GeneMapper ID Software v3 1 User Guide Using Panel Manager This chapter describes the Panel Manager window in the GeneMapper ID Software Version 3 1 Note For AmpF STR data analysis you use the Panel Manager only to import predefined panels and bins supplied by Applied Biosystems Most of the information provided in this chapter is not relevant to AmpF STR data analysis However this information is provided to explain various features and actions that you can perform using the Panel Manager for microsatellite and some SNP analysis For application specific tutorials refer to the Microsatellite Analysis with ABI PRISM GeneMapper Software v3 0 Tutorial PN 4335525 and the SNP Genotyping with ABI PRISM GeneMapper Software v3 0 Tutorial PN 4335524 This chapter covers Panel Manager Overview ceo a1 amp 6 806 s aera ewe hee eee ae 4 2 Panel Manager Window Commands 00008 4 7 Formats of Panel and Bin Text Files 4 22 Panel Table View i428 wectiud E oie ee eee yaad cot ee iets eet 4 34 Marker Table View s acanexo6 sn ecard i ened we Ra aes 4 36 Bins View Microsatellites Only 000000000 4 39 GeneMapper ID Software v3 1 User Guide 4 1
216. he Sample Name Color Info and the Color Comment fields of a sample sheet into GeneMapper JD software Project tables It is your responsibility to ensure that the entries on the sample sheet for your particular ABI PRISM instrument are correctly set up to provide GeneMapper I D software with the necessary information GeneMapper ID Software v3 1 User Guide Information from the Data Collection Sample Sheet or Plate Record Sample Sheet Plate Editor File Edit Plate Name eiGieiaiis Comments GeneMapper ID Software Project Table ka r Baji E Project 4 JRun_dermo_ al 6 Ll Hod Table mi SEMOWHES Status Sample Name wy Sample Type Specimen Category Analysis Method Panel Size Standard T OOSO OCOCOCisN sOC tsC C User defined Read from sample file GeneMapper ID Software v3 1 User Guide G 3 Appendix G Sample Sheets and Plate Records G 4 Find Command You can increase the utility of the Find Ctrl F command in GeneMapper D software by carefully planning the format of the information you put into the Sample Info or Color Info field of the sample sheet Example of How To Use the Find Command One Method If Then you have 12 samples numbered 1 2 3 12 and you enter these numbers into the Sample Info field when you search for all dye lanes conta
217. he number of rows are more than what can be accommodated on a page then the printing goes beyond a page vertically Now if the table has both the situations meaning number of columns are 50 5 pages horizontally and number of rows are 1000 10 pages vertically the printing can be done in 2 ways total 50 pages Either the pages can be printed horizontally first 5 pages with all columns and 100 rows or the pages can be printed vertically first 10 pages with all rows and 10 columns GeneMapper ID Software v3 1 User Guide Page Set Up Options e Prints Columns First This option completes printing all columns for each page of samples before moving on the next page of samples see order below Columns 1 5 Columns 6 10 Columns 11 15 Samples 1 20 Samples 21 40 Page 1 Page 2 Page 3 Page 4 Page 5 Page 6 e Print Rows First This option completes printing all Samples rows first for each page before moving on to the columns see order below Columns 1 5 Columns 6 10 Columns 11 15 Samples 1 20 Samples 21 40 Page 1 Page 3 Page 5 Page 2 Page 4 Page 6 e Use Screen Font The users can change the fonts of system via the operating system If the fonts are changed and this option is selected during printing system fonts and not application fonts are used e Print All Data There is special handling for large data in a column For example
218. hen when you click the Show Project Alleles icon il the s will be overlaid by a set of symbols marked as blue on the monitor which represent the data associated with a particular bin for the current project in the Project window and called according to the allele associated with the bin Bins may be used for calling alleles with or without allele calls in the reference data but project alleles s are required for automatic bin generation the Auto Bin function Reference alleles s are identified as a result of the Auto Bin command Selecting a reference data sample in the navigation pane displays that particular sample s electropherogram with bins overlaid to provide easy viewing while editing markers X and Y axes Genotype Quality Symbols and Cursor Location Information The Bins view X axis is the fragment size in base pairs the Y axis is quality or signal height if a reference sample is selected Vertical shaded bars are the bins that will be used to call alleles for the selected marker The symbols blue stars described above as Allele Call indicators represent Genotype Quality values on the Y axis for the alleles of samples that have been analyzed and are currently displayed in the open project The equivalent positions in the reference data are marked with red s The Genotype Quality values representing analysis results are not editable Bin location boundaries and names are editable Cursor locati
219. ide Using GeneMapper Manager This chapter provides information on how to use the GeneMapper Manager window of the GeneMapper ID Software Version 3 1 This chapter covers roduc UO iks era eier aTe hoe So ie Bela ae ear 5 2 Section 5 1 Projects Tab ccc cece cece cece eter ccees 5 3 Section 5 2 Analysis Methods Tab ccc ccc eee eees 5 9 Section 5 3 Table Settings Tab cece eee eens 5 27 Section 5 4 Plot Settings Tab cc ccc cece ee wees 5 41 Section 5 5 Creating and Evaluating a Matrix 5 53 Section 5 6 Size Standards Tab ccc ce ee ee ee eee 5 65 GeneMapper ID Software v3 1 User Guide 5 1 Chapter 5 Using GeneMapper Manager Introduction The GeneMapper Manager is a centralized interface within the GeneMapper D software v3 1 used for managing and organizing projects and other various information Six tabs assist you in customizing your data analysis e Projects tab e Analysis Methods tab e Table Settings tab e Plot Settings tab e Matrices tab e Size Standards tab of Samples e Project 2003 06 12 16 30 35 amid E Cace work Project 1 2003 06 12 16 40 35 girre Casework Project 2 2003 06 18 13 30 41 grmic Casework Project 3 2003 06 18 13 33 3 omnia Rename Save AS Inport Export Delete Figure 5 1 Tabs of the GeneMapper Manager 5 2 GeneMapper ID Software v3 1 User Guide Section 5 1 Overview Sect
220. iew sample and genotype plots and sample and genotype table including all PQVs and allele calls 1 Make sure the GeneMapper database you are importing the project into contains the same exact panels bins analysis method and size standard originally used to create the project If this is not the case export these items from the original copy of GeneMapper and import them into the GeneMapper copy on which you would like to view the data Note This step must be done prior to importing the project 2 Export the project from the original GeneMapper database and import it into another GeneMapper database of choice These operations are done within the GeneMapper Manager window GeneMapper ID Software v3 1 User Guide Transferring Data 3 Open the imported project from the File menu in the main project window You now have access to your data and genotypes You can also make allele edits Please note that you will not have access to the raw or EPT data nor will you be able to reanalyze the data Therefore if you make a marker or bin modification you can save these changes but you will not be able to reanalyze using those changes Refer to the next procedure to gain access the raw and EPT data and for reanalysis capabilities To view raw data EPT data and to reanalyze imported projects in addition to the above Note When you first analyze a set of samples and create a project a link 1s formed between the sample file s o
221. igation of a ROX dye labeled 22 mer oligodeoxynucleotide to the cut ends A subsequent enzymatic digestion with BstUI yields DNA fragments containing a single ROX dye See GeneScan 350 Molecular Lengths below The following table lists the GeneScan 350 Denatured Fragment Molecular Lengths Nucleotides for the 12 fragments Fragment Fragment Fragment Length Length Length 35 139 250 50 150 300 75 160 340 100 200 350 The following screen shows an electropherogram of GeneScan 350 run under denaturing conditions Size Match Editor Size Match Quality 0 99 439 150 160 2235 2465 2695 2925 3155 3385 3615 3845 4075 4305 4535 4765 4995 5225 5455 5685 3915 6145 6375 6605 GeneMapper ID Software v3 1 User Guide GeneScan 400HD Size Standard GeneScan 400HD Size Standard About This Size Standard Special Uses How It Is Prepared Fragment Lengths Denaturing Fragments The GeneScan 400HD High Density Size Standard is used to determine fragment lengths between 50 and 400 base pairs The high density of marker bands in this standard makes it particularly useful for microsatellite analysis All fragments have been checked for migration that is true to size under a wide variety of run conditions on all ABI PRISM instruments All aspects of the preparation of the GeneScan 400HD Size Standard are proprietary Each fragment c
222. indow size value B 19 PQV results A 5 Preferences Analysis tab 6 7 C 22 Import tab 6 5 C 20 Users tab 6 9 C 24 Print dialog box C 33 printable views 10 2 printing genotype plots 10 8 page options Plot Tab 10 6 page options Table Tab 10 4 sizing plots 10 7 printing considerations 10 2 Process Component Quality Values PQV 1 4 process quality values PQV editing A 2 overview A 2 PQV flags on the Genotypes page A 6 PQV flags on the Samples page A 5 PQV values for specific applications A 4 process A 2 rules for PQV columns A 5 using the PQV system A 3 Project Analysis menu commands C 13 Project Edit menu commands C 12 Project Edit menu dialog boxes Find dialog C 45 Sort dialog C 46 Project File menu commands C 10 Project File menu dialog boxes Export Table dialog C 31 New Project C 27 Open Project dialog C 28 Print dialog C 33 Save Project Save Project As dialogs C 29 Project Help menu commands C 26 Project View menu commands C 15 GeneMapper ID Software v3 1 User Guide Project window importance of 3 2 interface conventions 3 3 navigating in 3 12 parts of the main window 3 4 reformatting 3 3 tasks performed in 3 3 toolbar 3 18 viewing contents 3 14 projects workflow for storing K 1 Q Quality Flags tab options 5 25 PQV thresholds 5 24 5 25 R Raw Data tab 3 7 registration card completing 2 6 S sample files workflow for storing K 1 Sample Information source of G 2
223. ing Adobe Acrobat Reader About GeneMapper ID Opens the About GeneMapper ID window that indicates registration information and software version number 8 28 GeneMapper ID Software v3 1 User Guide Exporting Table Data This chapter describes how to export table data This chapter covers IN FOGUCHON ite Sees head ation dad ei teenie hdes 9 2 Exporting Samples and Genotypes Table 9 2 ADO CODIS erresa Boateng ate aed bees dee diets eS at Se 9 5 CODIS Export Manacer lt 2 6e 4o458 dae tbh aS 2Ghs EEEa 9 6 CODIS Table PXpOrt 3 322040 ctader ee tines ae ade a2 aE 9 8 GeneMapper ID Software v3 1 User Guide 9 1 Chapter 9 Exporting Table Data Introduction You can export the samples table or the genotypes table These tables can then be imported into a spreadsheet application such as Microsoft Excel In addition the data can be exported in a compatible FBI Laboratory Combined DNA Index System CODIS format Exporting Samples and Genotypes Table Viewing the Exported Table in a Spreadsheet Application 9 2 The exported table reflects what is displayed in the samples or genotypes tabs Therefore the exported table depends on what table setting you have selected when you export the table In addition the table is organized in a vertical format displayed one marker per row The sample and genotype PQVs do not display as color symbols when you import the table into
224. ing the full name of the column the default names are often acronyms e PQV results are reported in the column labeled SQ on the Samples page and GQ on the Genotypes page e The SQ and GQ quality indicators displayed in either symbols or in numerical values depending upon the Quality Metrics Display setting in the Tools Options Analysis tab have the meanings listed below Green good data default numerical range of 0 75 to 1 0 Yellow questionable data default numerical range of 0 25 to 0 75 Red low quality data default numerical range of 0 0 to 0 25 PQV Flags onthe The PQV flags on the Samples page Samples Page PQV Column Description SFNF Sample File Not Found e A flag Pass or Check A is automatically displayed when no sample file is found e Not editable MNF Matrix Not Found e A flag Pass Ii or Check A is automatically displayed when no matrix file is found attached to the sample file e Not editable SNF Size Standard Not Found e A flag Pass or Check A is automatically displayed when no size standard is found e Not editable GeneMapper ID Software v3 1 User Guide A 5 Appendix A Process Quality Values PQV Column Description OS Offscale e A flag Pass Ii or Check A is automatically displayed when there is offscale data in the signal e Not editable e ndicates an offscale result in the size standard region of the signal
225. ingle selection only Size Calling curve Displays the size calling curve for best fit OK button Closes the Size Match Editor window accepting the pending changes For samples whose size standards are modified the Project window is ready for reanalysis Cancel button Closes the Size Match Editor window disregarding any pending changes Apply button Same as OK button except the dialog box remains open Override SQ Use to override the Sizing Quality value and button automatically set the value to 1 0 This button overrides the sizing quality A check mark displays in the SQ column in the Project window Samples tab GeneMapper ID Software v3 1 User Guide 15 Chapter 7 Analyzing and Interpreting Data Editor The following Size Match Editor menus are described in this section Commands lt Edit e View e Tools Edit Menu The Edit menu is used to delete or change size labels View Tools el piel ei a ae Aud eee abel Delete Size Label Delete Change size Label Delete All Size Labels Ctrl Delete Edit menu items and descriptions ltem Description Enabling Undo Undoes the last user action Enabled after a user Ctrl Z action that adds modifies or deletes Add Size Adds a size label to the Enabled when an Label selected peak unlabeled peak is selected Delete Size Deletes the size label of the Enabled when a Label selected peak labeled
226. ining a 1 in the Sample Info field not only will you select sample 1 you will also get samples 10 11 and 12 A Better Plan If Then you number the samples 01 02 03 and so on a search for the text 01 would select only the desired dye lanes GeneMapper ID Software v3 1 User Guide Troubleshooting Sheet H This appendix covers Troubleshooting Sheets nananana anaana aaa GeneMapper ID Software v3 1 User Guide Appendix H Troubleshooting Sheet Troubleshooting Sheets Due to the complex nature of troubleshooting these following matrices are provided as a tool to collate the variables involved in the troubleshooting process with support groups product groups and the software development team at Applied Biosystems To address any issues that may arise when installing or running the software please fill out the appropriate list of questions and collect the necessary files before contacting Technical Support For Installation Collect the following files and complete the questions below Issues e install log X AppliedBiosystems GeneMapper e GeneMapper Log txt X AppliedBiosystems GeneMapper Does the computer and operating system meet the required computer specifications List any potential discrepancies Did you receive any error messages or alerts If so what did they say if possible take screen captures At what point during the installation process did the
227. installation TCP IP must be installed prior to Oracle installation 2 2 In order to perform all functions in GeneMapper JD software with its embedded Oracle database ensure there are at least 500 MB of free space on the partition where the GeneMapper D database is installed It is possible to export your projects to a different drive or partition then delete those projects from the database to make room for new projects See Chapter 3 Using the Project Window GeneMapper ID Software v3 1 User Guide Installing the GeneMapper ID Software Installing the GeneMapper D Software Compatibility GeneMapper ID software uses an Oracle database GeneMapper ID with Oracle software v3 1 is compatible only with the Oracle database installed Databases with ABI PRISM 3100 3100 Avant Data Collection software v2 0 Special The following is a list of special considerations for installing the Considerations GeneMapper ID software v3 1 e Compatibility of GeneMapper JD Software v3 1 with ABI PRISM 3100 3100 Avant Genetic Analyzer systems 3100 Data 3100 Avant f Collection Data Collection Compatible with x Instrument Computer Version Version 1 1 No use a separate computer that meets the 1 0 minimum requirementsa 2 0 2 0 Yes a GeneMapper ID software v3 1 cannot exist with other Oracle servers except the Oracle database installed by 3100 3100 Avant Data Collection version 2 0
228. ion 5 1 Projects Tab Projects Tab The GeneMapper Manager Projects page shown below is a utility used to rename save as import export or delete projects To display the Projects tab select Tools gt GeneMapper Manager gt Projects F GeneMapper Manager Projects Analysis Methods Table Settings Piot Settings Matrices Size Standards Project Last Saved Owner of Samples can Project 2003 08 1 216 30 3 gmc 8 asework Project 1 2003 06 12 16 40 3 gmid Casework Project 2 2003 06 18 13 30 41 amid Casework Project 3 2003 06 15 13 33 3 arrid Rename Save Ag Import Export Delete Figure 5 2 Projects Tab of the GeneMapper Manager Select the project you would like to change and click the appropriate button If you click the Delete button the GeneMapper Deletion Alert box displays Click No if you do not want to delete the project Click Yes only if you want to delete the project permanently GeneMapper Deletion Alert Ea 2 Delete Casework Project This wil permanently remove the object tram the database and cannot be undone Figure 5 3 GeneMapper Deletions Alert Box GeneMapper ID Software v3 1 User Guide 5 3 Chapter 5 Using GeneMapper Manager 5 4 Projects Tab Table 5 1 Elements Description of elements on the Projects tab Item Description Project column Project name Last Saved column Owner column Date Time stamp showing when the
229. iption GQ continued Genotype Quality Mathematical Formula e The mathematical formula used is AJA GR2119 5 5 X AQ GQ MQ x PQV x POV x xPQV a AQ Allele Quality is a function of quality value assignments for oF the following sizing quality allele calling quality bin assignment quality and bin quality MQ Marker Quality is modified by user defined PQVs to generate the final GQ value PQVs are weighted from 0 to 1 The actual value of a PQV in the equation is 1 minus the weight O weight no effect on the final GQ calculation 1 minus 0 1 therefore no change to GQ value 1 weight full effect 1 minus 1 0 If you multiply GQ by 0 and you get a GQ value of 0 then the sample analysis is of low quality Between 0 and 1 the higher the value the greater the impact on GQ PQV filtering is controlled by the threshold set by GeneMapper D software users and remains fully functional irrespective of what weight is chosen GeneMapper ID Software v3 1 User Guide Software Genotyping Algorithms This appendix provides a description of the new genotyping algorithms used in the GeneMapper ID Software version 3 1 and a detailed description of the Peak Detection Basic Classic and Advanced algorithm settings This appendix covers GeneMapper ID Software Genotyping Algorithms B 2 Basic Mode Peak Detection Algorithm Settings B
230. iption to a new profile Note You cannot change the name of an existing profile only the description Table Setting Editor New Profile Figure 5 18 General Tab of the Table Settings Editor 5 30 GeneMapper ID Software v3 1 User Guide Section 5 3 Table Settings Tab The Samples Tab The Samples tab provides the capability to filter columns in the Samples table of the project window Table Setting Editor Sho UE EE Show All Records N A E Semeer stow anRecores Sameie Name show anRecoras Figure 5 19 Samples Tab of the Table Settings Editor The Genotypes The Genotypes tab provides the capability to filter columns in the Tab Genotypes table of the project window and the capability to control the allele settings d Table Setting Editor a i 35 e User Defined Show All Records User Defined Show All Records Sow nee _ Figure 5 20 Genotypes Tab of the Table Settings Editor GeneMapper ID Software v3 1 User Guide 5 31 Chapter 5 Using GeneMapper Manager The Sample and Genotypes Tab 5 32 Elements The Samples and Genotypes tabs display the Table Setting properties that will be applied to the corresponding tabs in the Project window The properties or elements are described in the table below Samples and Genotypes Settings Table 5 8 Elements and settings for samples and genotypes data Element Description Column settin
231. ir range and dye color that define an allele A set of bin definitions for one source or set of experimental conditions usually an instrument bin sets are available inside a kit Having 2 or more alleles respectively per gene or locus The set of allele calls for a marker or genetic locus usually 2 alleles GeneMapper ID software Human identification and forensic applications A set of panels the grouping of panels in a kit is determined by the kit provider A known microsatellite or SNP genotyping location Short tandem repeat marker di tri tetra nucleotide repeat A set of bin definitions for one or more markers the grouping of markers in panels is determined by the kit provider Process Quality Value Quality values assigned to bins based on the success rate of the bins to locate allele peaks correctly Primer extension based chemistry for SNP genotyping validation Single nucleotide polymorphism used in this document to refer to SNaPshot Kit System markers Glossary 1 Glossary 2 GeneMapper ID Software v3 1 User Guide Index A Add Reference Data command 4 17 Add Sample to Project dialog box C 37 Add Samples dialog box Edit Menu C 38 Files tab C 41 View Menu C 40 Add samples to the Project C 38 advanced mode peak detection options B 15 ranges options B 12 size calling method options B 14 smoothing and baselining options B 13 algorithms allele calling B 2 B 3 binning B 2 B 3 peak detection B 2
232. is is a display filter for files folders are always shown Use to select the file extensions for display e All Files shows all files e Java Serialized file ser shows only files with the extension ser which is how GeneMapper D database projects are exported Note Only one preset file extension ser can be used Table 5 3 ltem Name 1 Toolbar 2 Drop down directory menu 3 Folder file display pane 4 Import button 5 File name field 6 Files of type field 7 Cancel button Closes the Import Project window without importing a project 5 8 GeneMapper ID Software v3 1 User Guide Section 5 2 Section 5 2 Analysis Methods Tab Analysis Methods Tab Overview The Analysis Methods Tab is used to create custom analysis methods for analyzing sample files An analysis method is a set of algorithm parameters that are applied to data during analysis For additional information about algorithms see Appendix B Software Genotyping Algorithms The Analysis Methods tab contains a list of analysis methods in the database To display the Analysis Methods tab select Tools gt GeneMapper Manager gt Analysis Methods a GeneMapper Manager Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Name Last Saved Owner Instrument Analysis Type Description HID Advanced 2003 06 12 14 09 71 gmid 37r analyzer HID HID_ Classic 2003 06 12 14 0
233. is of low quality You can set the threshold for the GQ PQV values in the Analysis Method Editor on the Quality Flags page GeneMapper ID Software v3 1 User Guide A 3 Appendix A Process Quality Values SO l JOPPE 10 JON GeneMapper ID Software v3 1 User Guide 19410 TESTAN Se ee S N emerson apHosjonu ip p hi hiM D PIPER MER A AER ER NEM A N N emesan S NINININININ ININ ININA N NIN ens NININA NINJA MERI AID IA S ih Rh ah OS 4NSS 4NW 4N4S dd 9N 740 99 SO dS d9 NdS Hd1 YHd NIJ WdS vaO NV dHS av oav PAIEA AOd ofqvordde joN yew yooyo ON SISATBUR sry YIM posN N OI IICSOIDT MYA glOYSd NS surdAjousy ANS ANS pue uoneoyyuopg uewny AIH SISA BUL SUIMOT OJ OY JO YORO OF posn OJE SON VA YOIYM SMOYS Qe SUIMOT OJ OY SosATBU orroads IOF pasn oe sonje A d ULL suoneoyddy oads 10 SonjeA AOd lt 4 Using Process Component Based Quality Values Rules for PQV In gaining familiarity with the PQV columns on the Samples page Columns and Genotypes page keep the following rules in mind e Quality metrics with Pass Check values and no Low Quality value are warning flags Analysis does not stop if problems are detected with these properties but you should examine results flagged with Check values e Holding the cursor over a column header displays a ToolTip identify
234. is range to start after the primer peak Note Sample files generated from the ABI PRISM 377 DNA Sequencer may have already removed the primer peak by setting the data point range for gel image generation to exclude the primer peak Sizing Enter Start and Stop size numbers in the entry drop down menu fields in order to specify only a limited range All Sizes analyzed for each sample Partial Sizes GeneMapper ID Software v3 1 User Guide B 7 Appendix B Software Genotyping Algorithms Data Processing The Data Processing parameter options specify how the raw data is processed before peak detection and size calling B 8 Parameter Options Item Description Baseline Used to automatically adjust the baselines of all checkbox detected dye colors to the same level for a better comparison of relative signal intensity Multicomponent checkbox Note Do not check this box for multicapillary instruments This user function was performed in Data Used to specify that the GeneMapper ID software applies a predefined matrix to adjust for spectral overlap when it performs analysis Although the dyes used to label DNA fluoresce at different wavelengths the spectra overlap to some extent Create a matrix file containing a mathematical matrix to correct for this overlap For a description of matrix files and how to create option buttons Collection them see Section 5 5 Creating and Ev
235. itor The Size Match Editor toolbar contains the following icons Icon Description Enabling L the size of other peaks automatically tothe peak is Autoadjusts size standard peaks Adjusts Enabled after a right of the selected peak a changed Tooltip Auto Adjust Sizes Checks the quality of the current size Enabled after a e matching Checks the quality of the peak label is revised peak positions and updates the changed Size Match Quality indicator Tooltip Check Sizing Quality Deletes all size labels Always Pai enabled Tooltip Delete All Size Labels a Only for Basic and Advanced modes The Size Match Editor window uses the mouse for editing Note All user edited size standard samples automatically have a sizing quality of 1 0 and display a check mark in the SQI column in the Samples window To edit a particular size standard peak l 2 Click the peak to be edited Right click the peak to open the editing pop up window and Add Delete or Change the selected peak If you chose Add or Change move the cursor to the right to open the Select Size sub menu Choose the desired height value from this menu Check the sizing quality by selecting Check Sizing Quality under the Tools menu If the peaks are sized correctly but the quality score is still below passing click the Override SQ button to set the SQ to 1 0 Click Apply if you are going to edit a second size
236. itor window controls how the Allele Size Height Area Mutation and Comment columns are displayed in the Genotypes view Allele Settings Mumber of Alleles F Keep Allele Size Height Area Mutation and Comment together cancel Table 5 9 Allele settings and descriptions Allele Settings Description Number of Alleles The number of alleles per marker that are displayed in the genotypes table The default value is 2 However when you are analyzing mixtures or want to have more than two alleles per marker in the table you need to increase this value Keep Allele Size Height Area Mutation and Comment together If the box is unchecked then the columns display as Allele 1 Allele 2 Size 1 Size 2 Height 1 Height 2 Area 1 Area 2 Mutation 1 Mutation 2 Comment 1 Comment 2 or If the box is checked then the columns display as Allele 1 Size 1 Height 1 Area 1 Mutation 1 Comment 1 Allele 2 Size 2 Height 2 Area 2 Mutation 2 Comment 2 GeneMapper ID Software v3 1 User Guide 5 33 Chapter 5 Using GeneMapper Manager Creating a New To create a New Table Setting Table Setting l Select GeneMapper Manager gt Table Settings and click New to open the Table Setting Editor window In the General tab enter a name for the new setting and a description if necessary Open the Samples and or Genotypes tabs and select the desired contents for the Samples and
237. izing Partial Range Partial Sizes Start Ft 2700 Start Size 75 Stop Pt reo Stop Size 450 Smoothing and Bazelining Peak Amplitude Thresholds Bf e fs G 50 0 fo Y fo Min Peak Half Wwicdth 2 pits Polynomial Degree 3 Peak Window Size is pts Slope Threshold Peak Start bo Peak End joo Factory Defaults Smoothing i None i Light C Heavy Baseline Window sr pits Size Calling Method 2nd Order Least Squares 3rd Order Least Squares Cubic Spline Interpolation Local Southern Method C Global Southern Method Figure 5 12 Peak Detector Tab of the Analysis Method Editor Advanced Peak Detection Algorithm GeneMapper ID Software v3 1 User Guide 5 19 Chapter 5 Using GeneMapper Manager 5 20 e Analysis Range defined in scans not by size Full Range the software analyzes the raw data in the entire scan region as collected by the genetic analysis instrument including the primer peak Partial Range the software analyzes data between the user defined scan points When this option is selected Applied Biosystems recommends that you select a Start Point after the primer peak and before the first size standard peak you want to define The Stop Point should be after the last size standard fragment you define This range may vary from instrument to instrument and platform to platform Sizing Range in base pairs Full Range the software reports all peaks within the range
238. l COfiler_vl Size Standard HID_Classic_G5500 250 Matrix STR Class Matrix 3 1 99 Analysis Method HID Classic Peak Detection Mode Classic Analysis range data point start 3275 stop 6400 Sizing range bp start 75 stop 400 Smoothing light smoothing Peak thresholds rfuj Blue 50 Green 50 Yellow 50 Red 50 Orange 50 Peak min width bp 3 Peak correction method none peak correction limit data point 30 Size calling method local Southern Run Information User Name FDLE Instrument ID ABI PRISM 310 Data Collection Ver ABI PRISM 310 Collection 1 0 4 Run Date Time 0002 11 30 00 00 00 0 Run Duration NA Total Data Points 6496 Data Collection Settings Module File GS STR POP4 1 mL F Run Voltage 15000 Injection Voltage 15000 Injection Duration 5 Temperature 60 Laser Power 9 J Ste 3 6 GeneMapper ID Software v3 1 User Guide Overview of the Project Window The Info tab displays sample file information such as e sample information e run information e data collection settings e gel information e capillary information e error messages e current settings Sample information comes directly from the Instrument Data Collection and or Sample sheet Raw Data Tab Clicking the Raw Data tab in the Samples view when the Sample file is selected displays the Raw Data view for the selected Sample file GeneMapper ID 3 1 Casework Project gmid Is Logge
239. l 5 Appendix GeneMapper ID Software Verification Instrument Table I 2 lists the instrument platforms and computer operating Platform and systems that were used to electrophorese the samples for verification Computer testing eee Table l 2 Instrument and computer platforms Instrument Computer Operating System 310 Genetic Analyzer Apple Macintosh v9 1 Microsoft Windows NT 377 DNA Sequencer Macintosh v9 1 Microsoft Windows NT 3100 Genetic Analyzer Microsoft Windows NT 3100 Avant Genetic Analyzer Analysis Software The software packages and computer configurations used to analyze the data are listed below Table l 3 Software and computer configuration used for analysis Genotyper Software v2 5 2 Analysis Software and Version Computer Configuration Color Printer GeneScan Software v3 1 2 Macintosh G3 128 MB RAM v9 1 HP LaserJet operating system 4500N GeneScan Software v3 7 1 Genotyper Software v3 7 GeneMapper D Software v3 1 Classic and Advanced modes Dell Latitude C810 with a 1 1GHz processor 512 MB RAM Microsoft Windows 2000 operating system Service Pack 3 l 6 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Results of Verification Testing The verification was designed to evaluate four specific categories e Peak detection and genotyping e Algorithm testing e Workflow e Data handling
240. lal Fla lia ii EE P OT P Sample Name Pand Marker 0 S BIN PHR LPH SPU AN BD CC OVL GQ Samples Identifile_v1 cro D ry O EHM E amp X 310 64 Y 1918 Figure l 20 The default peak height ratio value is set at 70 and the maximum allele number is set at 2 in GeneMapper ID software Any peak below these values activates the PQV flags The ratio between the first and second peak is 64 and the ratio between the first and third peak is 36 resulting in a yellow peak height ratio PQV flag Low Peak Height The tenth parameter tested in the Algorithm Testing category is the PQV Flag individual PQV for the low peak height LPH flag verification using both the Classic and Advanced modes 2 772 alleles from low quantity DNA samples and 16 alleles from a single source DNA sample were evaluated Results 100 of the alleles tested accurately reflect the LPH PQV l 30 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Control The eleventh parameter tested in the Algorithm Testing category is Concordance the individual PQV for the control concordance CC verification PQV Flag using both the Classic and Advanced modes 198 alleles for Control DNA 9947A 96 alleles for Control DNA 007 and 30 negative control amplifications were evaluated Results 100 of the alleles tested accurately reflect the CC PQV Overall GQ PQV The twelth parameter tested in the Algorithm testing category is the fi
241. le 5z2 ing 4 Yelow Female 52 ing 1500 1000 00 a m ne 1705 1683 1647 1949 1442 1125 44 Female_a2_ing 4Red Female_a2_ing 2000 2000 1000 11 18l z224 1797 1629 1673 Figure l 1 Female single source DNA Sample amplified with the AmpF STR Identifiler PCR Amplification kit electrophoresed on a 310 Genetic Analyzer Macintosh OS then analyzed with GeneScan Software v3 1 2 and Genotyper v2 5 2 l 10 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Samples Plot ey lal x File Edit View Tools Alleles Help Plot Setting HID_Classic gt E Panes fa TT a EP al l Vi xf fe LPF Sample File Sample Name Pand sgo SQ here are no controls in this selection Ad Female_32_Ing fsa Fenale_32_Ing Ideniifile_v1 x i Ds S De A O GSI D7 820 110 120 130 140 150 160 170 180 1390 200 210 220 230 240 250 260 270 280 230 300 310 320 330 340 350 360 1200 1582 1589 1098 1500 1353 1082 968 Ad Fanale _32_Ing fsa Female 32_Ing Wdentifiler_v1 x gO D351358 DBS Z JDE D251338 110 120 130 140 150 160 470 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 0 290 300 310 320 330 340 350 360 Ad Female 32_Ing fsa Female 32_Ing Tdentifiter vl x O D55818 ECE 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 xX
242. le Selected for G Data start At 1000 uP Mo File Selected for Y Data Start At 1000 k Mo File Selected for R Data Start At 1000 a Mo File Selected tor O Data Start At 1000 Points 100000 e vy r Figure l 12 Matrix created in the GeneMapper ID software using the Advanced algorithm Size Standard The sixth parameter evaluated in the Peak Detection and Genotyping Test category is the size standard test using the size match editor in the Classic and Advanced modes The size match editor calculates size standards differently depending on the algorithm used to generate the data e Classic algorithm If the Classic algorithm is used to analyze data a size quality value of 0 5 is associated with each sample Figure I 13 on page I 22 The size standard peaks are identified by the manual assignment of size standard definitions to the associated data points within a run similar to GeneScan software v 3 1 2 Macintosh OS GeneMapper ID Software v3 1 User Guide l 21 Appendix GeneMapper ID Software Verification e Advanced algorithm The Advanced algorithm however has a size quality value ranging from 0 0 to 1 0 associated with each sample Figure I 14 on page I 23 Ratio matching is used to identify the size standard fragments similar to GeneScan software v3 7 1 Windows NT OS You can check and edit the sizing result The sizing result is applied to the final genotype process component quali
243. le na export HD Ch To see the Click the And GeneMapper ID software displays the Genotypes view Genotypes tab Project in the Genotypes view Alternatives GeneMapper ID v3 1 Casework Project gmid Is Logged In N t S Select File Edt Analysis View Tools Help ote Samples ce See Se ae eel e z mustbe PSS Genotypes Ae Aee E rlo Tae analyzed to see commane from wa Samples Genotypes genotypes the View menu E cOrfier v1 Sample Mare Run Mare Panel hal i or Ctrl Shift 2 Profiler Pius_vt h cO _Cortrol Casework COfiler_v1 Da 2 CO_Contral Casework cofiler_v1 Dt 3 CO_Contral Casework Eiler v E 4 CO_Contral Casework cofiler_v1 TH To see the Click the And GeneMapper ID software displays the Samples view Samples tab Project in the Samples view again Alternatives GeneMapper ID 3 1 Database Project gmid Is Logged In Select View gt Ble Eo Arales mie loos HER Samples or E e aje gM MA a E BB T seting Ho Tae Ctrl Shift 1 e Coen Samples Genotypes H E Jbatabasingl Status Sample F Sample N Sample IC Comment aS T Specir o mee pe a no ex C E no ex ID_Sampl Sampled More Sample mo exp 3 14 GeneMapper ID Software v3 1 User Guide Navigating in the Project Window Access to The following table lists how to access subordinate GeneMapper ID Subordinate software windows Application Windows To see the
244. lect View gt Raw Data to display the raw data Then select File gt Print Note You cannot print any displays from within the Panel Manager Hard copies from the Panel Manager can be obtained by taking a screen shot of the desired view then printing the screen shot Printing Things to consider when printing Considerations e The more information displayed in a printout the slower the page s print To maximize printing speed configure your displays with the minimal amount of data needed For example the printing speed is faster if you label your peaks with one label each instead of four labels each e Ifyou do not need printouts to contain as much information as when you are editing and interpreting the data you can create plot settings specifically for printing within the GeneMapper Manager e The printout resembles the display on the screen If you zoom the printout shows the zoomed in area However if you are printing the sample or genotypes table and you have adjusted the column widths the adjusted widths are not displayed in the printouts e When printing the samples or genotypes tables usually all columns do not fit on one page Refer Page Set Up Options on page 10 4 for more information 10 2 GeneMapper ID Software v3 1 User Guide Printing Data e You can export these tables and adjust the column widths within a spreadsheet application such as Microsoft Excel to fit on one page e The speed of prin
245. lect a sample all of the corresponding genotypes are automatically selected in the Project Window Genotypes Tab If a plot window is open the view refreshes as you switch between tabs GeneMapper ID Software v3 1 User Guide About the Plot Window When you select a genotype the corresponding sample s are automatically selected in the Project Window Samples Tab If a plot window is open the view refreshes as you switch between tabs The genotype selection is maintained until additional samples or genotypes are selected or deselected Note Samples with a low sizing quality SQ value cannot be displayed in the Plot window Only raw data can be viewed for low quality samples Note The Plot Window cannot contain electropherograms from both the Samples and Genotypes tabs at the same time Customizing Plot You can customize certain features within the Plot Windows using Windows the pull down menu items or toolbar icons Frequently used plot views can be saved as Plot Setting profiles using the Plot Settings Editor accessible from the GeneMapper Manager e The last profile used in the Samples Plot is used to open the Genotype plot e The last profile used in the Genotypes Plot is used to open the Sample Plot GeneMapper ID Software v3 1 User Guide 8 3 Chapter 8 Using Plot Windows Samples and Genotypes Plot Window Toolbars Toolbar The toolbar icons enable and disable plot features similar to their Descriptions associated
246. ler vi 0351358 blue 98 00 148 00 14 15 4 0 110 none 12 13 14 15 16 17 18 19 VWA blue 151 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none 18 19 20 21 22 23 24 25 26 26 2 27 28 29 30 AMEL green 106 00 114 00 x 9 0 000 none KY THOL green 159 00 205 00 8 939 3 4 0 060 none 5 6 7 8 9 9 3 10 TPOX green 212 00 254 00 8 4 0 060 none Oy 7 8 9 10 11 12 13 CSF1LPO green 275 00 323 00 10 12 4 0 090 none 6 7 8 9 10 11 12 13 14 15 D55818 yellow 128 00 180 00 11 4 0 100 none 7 8 9 10 11 12 13 14 15 16 D135317 yellow 192 00 242 00 11 4 0 100 none 8 9 10 11 12 13 14 15 D75820 yellow 251 00 298 50 10 11 4 0 090 none Oy 7 8 9 10 11 12 13 14 15 Panel Profi ler_Plus_v1 D351358 blue 98 00 148 00 14 15 4 0 110 none gee Fei We Ie ee rg a VWA blue 151 00 203 00 17 18 4 0 110 none 11 12 13 14 15 16 17 18 19 20 21 FGA blue 206 25 360 00 23 24 4 0 110 none 18 19 20 21 22 23 24 25 26 26 2 27 28 29 30 AMEL green 106 00 114 00 x 9 0 000 none D8S1179 green 118 00 183 50 13 4 0 120 none Op 9 10 11 12 13 14 15 16 17 18 19 D21511 green 184 50 247 50 30 4 0 130 none 24 e gp eye Oe el poy 2Bs Py Os 2 Oe oy Sy Sle ey Bly Sleds Ses 52 D18551 green 264 49 350 00 15 19 4 0 160 none 9 10 L0 2 11 12 13 13 2 24 14 2 15 16 17 18 15 20 2 D55818 yellow 128 00 180 00 11 4 0 100 none 7 8 9 10 11 12 13 14 15 16 D135317 yellow 192 00 242 00 11 4 0 100 none Bp 9 10 11 12 13 14 15
247. les tab and samples are present Does not work with HID samples View Menu The View menu is used to hide show the Project window navigation pane and switch between the two Project window tabs The following table is provided as a reference for the View menu Tools Help Samples Ctrl Shitt 1 Ei peS Gif heifers Sample Info Ctrl F 1 Raw Data Ctrl F 2 EFT Data Ctrl F 3 BUIEN EY irl ne Stale show Mavigator Ctri Shift N Item Description Enabling Samples Switches to the Samples tab Always enabled Ctrl Shift 1 Genotypes Switches to the Genotypes Enabled when results Ctrl Shift 2 tab are present GeneMapper ID Software v3 1 User Guide C 15 Appendix C Project Window Software Interface Item Description Enabling Sample Info Switches to the Info tab for Always enabled Ctrl F 1 the selected sample in the project Raw Data Switches to the Raw Data Always enabled Ctrl F2 tab for the selected sample in the project EPT Data Switches to the EPT tab for Always enabled Ctrl F3 the selected sample in the project Full View Shows the full view of the Enabled only when Ctrl Raw Data or EPT graph viewing the Raw Data or EPT Data of a sample Y Axis Scale Changes Y axis scale See Enabled only when Y Axis Scale on viewing the Raw Data page C 16 or EPT Data of a sample Show Switches the navigator pane Always enabled Navigator shows hides the pane
248. lightly larger than the largest split observed This value set in data points specifies the maximum width of split that should be corrected the difference in data points of the positions of the two peaks GeneMapper ID Software v3 1 User Guide Appendix B Software Genotyping Algorithms Advanced Mode Peak Detection Algorithm Settings Overview In Advanced mode there are four main analysis parameter options They are as follows e Ranges e Smoothing and Baselining e Size Calling Method e Peak Detection Ranges Options The following are the Analysis Range options Item Description Full Range button Use to analyze all the data collected on the genetic analysis instrument for each sample Partial Range Enter Start and Stop data point numbers in the Data Points entry fields in order to specify only a limited range button to be analyzed for each sample Data points affect what is displayed in the results Normally you set the analysis range to start after the primer peak Sizing Enter Start and Stop size numbers in the entry drop down menu fields in order to specify only a limited range All Sizes analyzed for each sample Partial Sizes B 12 GeneMapper ID Software v3 1 User Guide Advanced Mode Peak Detection Algorithm Settings Smoothing and The Smoothing and Baselining parameter options help to optimize Baselining peak size and eliminate noise from the baseline O
249. lot 0 x File Edit View Tools Alleles Help Plot Setting AmpFLSTR Sizing E Panes fa EE E a EA Pai EE LAE E Sample Name Pand SsQO SQ Samplel cofle vl x E a 100 200 300 400 1600 800 0 C 200 200 400 fd lihituel adiba hott li 0 Sampla Profiler_Plus_vl x E 100 200 200 400 2000 1000 0 Sample Profle Pus X 75 0 100 0 11409 139 0 2036 12492 150 0 12626 160 0 12776 x 173 79 Y 1837 7 24 GeneMapper ID Software v3 1 User Guide Viewing Sample Data Editing Allele Ifyou are going through the labeled plots and you find a mislabeled Calls and Viewing peak you can edit these allele calls in the plot windows When Allele History editing alleles the following rules apply If you delete a label you will be prompted to enter an allele edit comment but the comment is not displayed in the genotypes table To see the allele edit comment you must display the allele history If you add a label to a previously deleted label the allele edit comment is displayed in the genotype table and an X is placed in the AE allele edit column in the genotypes table If you change the label on a peak you will be prompted to enter an allele edit comment This allele edit comment is displayed in the genotypes table and in the allele history and an X is placed in the AE allele edit column in the genotypes table When an allele is edited all PQVs for that
250. lumn to open the selections 2 Expand the appropriate folder 3 Double click the appropriate panel 4 Click Edit gt Fill Down 3 10 GeneMapper ID Software v3 1 User Guide Overview of the Project Window Genotypes View The Genotypes tab becomes enabled only when the data is successfully sized and allele calls are made In addition only samples that are sized correctly are displayed in the Genotypes view table When you perform a sizing only application this tab is not active after sizing since there are no genotypes to display GeneMapper ID 3 1 Casework Project gmid Is Logged In File Edit Analysis View Tools Help ek ce Sy PA L OD Bl f E rete seting fHo Tate 088 E Panels Samples Genotypes Sample Name Run Name Panel Marker Allele 1 Allele 2 AE Comrr AE Comrr ADO AE 8 Profiler _Plus_v1 CO_Control Casework COfiler_v1 D3S1358 14 15 CO_Control Casework COfiler_v1 D16S539 11 12 CO_Control Casework COfiler_v1 AMEL CO_Control Casework COfiler_ 1 THO1 9 3 f D I CO_Control Casework COfiler_ 1 CSF1PO 12 CO_Control Casework COfiler_ 1 D75820 C adder C ws z FO OOOO aa a L E CO_Control Casework COfiler_v1 TPOX 1I LE IEG IN BRD ee The genotypes table in this view contains items such as allele calls genotype PQVs and overall genotyping quality This t
251. m 0 to 1 e The actual value of a PQV in the equation is 1 minus the weight e 0 weight no effect on the final GQ calculation 1 minus 0 1 therefore no change to GQ value e weight full effect 1 minus 1 0 If you multiply GQ by 0 and you get a GQ value of 0 then the sample analysis is of low quality e Between 0 and 1 the higher the value the greater the impact on GQ PQV filtering is controlled by the threshold set by GeneMapper ID software users and remains fully functional irrespective of what weight is chosen 1 MQ Marker Quality is modified by user defined PQVs to generate the final GQ value 2 AQ Allele Quality is a function of quality value assignments for the following sizing quality allele calling quality bin assignment quality and bin quality GeneMapper ID Software v3 1 User Guide B 5 Appendix B Software Genotyping Algorithms Basic Mode Peak Detection Algorithm Settings Overview The Basic peak detection algorithm is used as the default algorithm by GeneMapper ID software v3 1 Basic Mode uses the Local Southern size calling method which determines the sizes of fragments by using the reciprocal relationship between fragment length and mobility In Basic mode there are two analysis parameter options e Minimum Peak Height Automatic e Minimum Peak Height User specified rfu Minimum Peak The Minimum Peak Height group box provides two parameter Height options ltem De
252. m plot settings or imported the factory provided defaults you can simply switch back and forth between different views by selecting different plot settings from the drop down box in the sample plot window 7 22 GeneMapper ID Software v3 1 User Guide Viewing Sample Data Viewing Sample Data IMPORTANT If the size quality SQ is low the samples will not be labeled genotyped GeneMapper JD v3 1 allows you to fully customize your plot windows You can create plot settings to re use or you can change the plot view as you work GeneMapper D v3 1 comes with several default plot settings These need to be imported into the plot settings tab of the GeneMapper Manager The plot settings supplied include AmpFLSTR AmpF STR Genotyping displays labels on peaks similar to Genotyping Genotyper plots Samples Plot E i x File Edit view Tools Alleles Help Plot Setting AmpFLSTR Genotyping E Panes fa BE E E aa WE EG W Em MPF I a Sample Name sQgo SQ here are no controls in this selection Samplel Profiler _Phus_vl x a F ms SSS SSS SSS SSS SSS SSS SSS 90 Ed Es Samplel Profle _Phus v1 x C zA SSS SSS eo E E 70 30 250 izle xX 139 51 Y 1012 Bin 19 Marker D351358 GeneMapper ID Software v3 1 User Guide 7 23 Chapter 7 Analyzing and Interpreting Data AmpF STR Sizing AmpF STR Sizing displays sizing data similar to GeneScan plots and sizing table Samples P
253. me save Files of type ava serialized file ser Cancel Figure 5 4 Export Project Window Note The sample files analysis method size standard panel and bin set used to analyze a project are not exported with the projects GeneMapper ID Software v3 1 User Guide 5 5 Chapter 5 Using GeneMapper Manager Using the Export Project Window 5 6 The elements called out in the figure in Export Project Window on page 5 5 are described in the table below Table 5 2 Export Project Callouts ltem Name Description 1 Toolbar Click these icons to Move display up one level in main pane Move display to Home level in main pane This level is usually Profiles lt user gt te Create a new folder at the present fad directory level oe Present a list of the contents of the B B selected folder Present details of the selected E folder 2 Drop down Select drive letter and or folder directory menu 3 Folder file The contents of the folder selected in the display pane toolbar are displayed here 4 Save button Saves the exported project 5 File name field Enter a file name to use for saving your data 6 Files of type This field is a display filter for files folders are field always shown Use to select the file extensions for display e All Files shows all files e Java Serialized file ser shows only files with the extension ser which is how GeneMapper
254. mn header cell to highlight the entire column or highlight a portion of the column that you want to apply the same setting to then select Edit gt Fill Down 4 Inthe Size Standard column define the size standard GeneMapper ID 3 1 Untitled gmid Is Logged In 5 x File Edit Analysis View Tools Help S ce all EL E p Tae seting Ho Tane H h S Project Samples elk Genotypes H E casework ae p a a F noexport Mone me New Size Standard 3 npexport None None o amas 5 ho export Mone Mone et Fi BEO N 0y 7 8 GeneMapper ID Software v3 1 User Guide Analyzing Samples Files a Use the drop down list to select the appropriate size standard Note You have the option to define a new size standard from this drop down list b Click the column header cell to highlight the entire column or highlight a portion of the column that you want to apply the same setting to then select Edit gt Fill Down In the Matrix column select a matrix a Use the drop down list to select the appropriate matrix for your 310 or 377 instrument b Click the column header cell to highlight the entire column or highlight a portion of the column that you want to apply the same setting to then select Edit gt Fill Down Select the desired table setting profile from the drop down list in the main project window Note You can change this table setting after dat
255. mple Files for details Set up the Project window for analysis of a given set of data Import the fragment analysis sample files Select the appropriate analysis method Choose the appropriate panels Select the size standard Perform e Perform analysis and examine the results i i Analysis Initiate analysis Examine analysis results using the Plot window s Export results e Export results Set up table format for export using Table Settings Editor Export using Export Table command GeneMapper ID Software v3 1 User Guide 1 5 Chapter 1 About GeneMapper ID Software Organization of the User Guide Chapters This manual provides technical information to support the GeneMapper D software user The following chapters explain how to use the major application components e Chapter 2 Getting Started with GeneMapper ID Software describes hardware requirements and procedures for installing registering and logging in and out of the GeneMapper ID software v3 1 e Chapter 3 Using the Project Window describes how to use the main Project window and associated utility windows e Chapter 4 Using Panel Manager describes how to use the Panel Manager to manage all of the chemistry kits panels markers loci and bin definitions required for automated allele calling e Chapter 5 Using GeneMapper Manager describes how to use the Proj
256. n 5 1 Projects Tab 0d 042 5 os Se0e0es Ges ee es 5 3 Section 5 2 Analysis Methods Tab 00000 e eee 5 9 Section 5 3 Table Settings Tab 0 0 00 cee ees 5 27 Section 5 4 Plot Settings Tab 00 c eee ees 5 41 Section 5 5 Creating and Evaluating a Matrix 5 53 Section 5 6 Size Standards Tab 0 0 c eee es 5 65 Options Dialog Box HEVOOUCTIONN itech a e See ts ata eat nade Maes ore deleted 6 2 HID Analysis OOuONnS iyu 49 ict seea wd She Ke ea 6 3 SACD Va Oy ier nu acer aen SS td A eos a ee ae eae 6 4 AGG Samples lab iasiva a Pew take pee er Pe ae ws 6 5 ANALYSIS TaD a Gu Ree eke ek ee ea a ee es Se 6 7 Users he ee clash a a E E E rec A nt ce ees 6 9 Analyzing and Interpreting Data INTFOGUGCTION sacadat regn oe es bs Rea cob eed See ee ees 1 2 Considerations for HID Analysis 0 000 cee eee eee 7 2 Analyzing Samples Files 0 00000 ccc eee ees 7 3 Interpreting Sample PQVs Sizing Quality and Precision 7 11 SIZE WAC EGON sions diet eee tp acn ty Benne Gib eaa by date dese a 7 14 Size Calling Curve 0 ce ees 7 19 Viewing Sample Data 2 0 0 0 0c eee 7 23 GeneMapper ID Software v3 1 User Guide Chapter 8 Chapter 9 Chapter 10 Appendix A GeneMapper ID Software v3 1 User Guide Genotypes Table ss diwali bute dena ate dele dee aa ds oe eho oe 7 27 Interpreting Genotype PQVs and Genotype Quality 7 30
257. n pane 2 Select the Add Bin menu item or right click within the allele size range of the marker shown in the Panel Manager right hand window to select the New Bin option 3 Drag the length of the desired bin range with the mouse 4 Edit the bin information in the Add Bin dialog box that opens For SNP Genotyping data 1 Select a SNP genotyping panel in the Panel Manager navigation pane 2 Select a marker in the Plot tab 3 Select the Add Bin menu item or right click on a marker or bin in the Plot tab 4 Edit the SNP marker using the dialog box that opens GeneMapper ID Software v3 1 User Guide 4 13 Chapter 4 Using Panel Manager Delete Bin The Delete Bin command deletes a marker bin as follows For microsatellite data 1 Select a marker in the Panel Manager navigation pane 2 Select a bin to be deleted 3 Select the Delete Bin menu item or right click the bin and select the Delete Bin option For SNP Genotyping data 1 Select a SNP genotyping panel in the Panel Manager navigation pane 2 Select a bin to be deleted in the Plot tab 3 Select the Delete Bin menu item or right click the bin and select the Delete Bin option Edit Bin The Edit Bin command edits a marker bin as follows For microsatellite data 1 Select a marker in the Panel Manager navigation pane 2 Select a bin to be edited 3 Select the Edit Bin menu item or right click the bin and select the Edit Bin option 4
258. n set into Panel Manager before you select it from the menu For non AmpF STR analysis you create the bin set in Panel Manager before it appears as an option in the drop down menu Marker Repeat Type for AmpFZSTR and other microsatellites only As stated on the Allele tab view no entries are required for dinucleotide repeats because the application calculates them automatically Check the Use marker specific stutter ratio if available check box to insert stutter ratio information into the Panel Manager marker s table If you know the stutter ratio for an individual marker you can set this number under the Marker Specific Stutter column in the Panel Manager Checking this box on the Allele tab makes the algorithm use the stutter ratios you previously defined in Panel Manager and not those defined in the Stutter Ratio box and the Allele tab Different values can be used for each marker IMPORTANT Use the marker specific stutter ratio for AmpF STR kits These values are provided in panels and bins supplied by Applied Biosystems The Cutoff value ignores all peaks less than the cut off ratio of the largest peak in the allele size range The MinusA ratio and Stutter ratio refer to the ratios you are expecting For example if you expect a stutter percentage of 20 for your markers enter in 0 2 GeneMapper ID Software v3 1 User Guide 5 17 Chapter 5 Using GeneMapper Manager The following table pr
259. nal PQV for the overall genotype quality GQ verification using both the Classic and Advanced modes 20 364 alleles were evaluated and accuractly reflect the overall GQ PQV except when off scale data is evaluated The genotypes table below shows two low quantity DNA samples amplified with the AmpF STR Profiler Plus kit For illustration purposes two loci D13S317 and D7S820 with their respective PQVs are calculated on page I 32 GeneMapper ID Software v3 1 User Guide l 31 Appendix I 32 GeneMapper ID Software Verification GeneMapper ID 3 1 offscale gmid Is Logged In File Edit Analysis view Tools Help e S l S e All L OO EL E OR Tavie seting HO Tanie EEE J Panels Samples Genotypes Profiler_Plus ample NelRern Name a DOLA O Bh pep TET BD O Q low _input Profiler_plus_NT Profiler _Plus_v1 E El N El E E H 71 g low _input Profiler_plus_NT Profiler_Plus_v1 D21511 G H M Mm m a a B A lovy_input Profiler_plus_NT Profiler_Plus_v1 D3513568 B ical a fw E E E C E Ez lovy_input Profiler_plus_NT Profiler_Plus_v1 D55818 Y m E m m H E E a lovy_input Profiler_plus_NT Profiler_Plus_v1 D75820 E E H A is 8 iz A lovy_input Profiler_plus_NT Profiler_Plus_v1 D6S1179 G a i A A E B E E A lov
260. nd bin sets Bins Menu _ The Bins menu contains commands to edit markers and bins Bins View Edit SWE Marker Ctrl H Delete SWE Marker CT S hit Add Bin Ctrl B Edit Bin Ctrl H Delete Bin Ctrl D Hew Bin set Delete Bim set show Project Alleles add Reference Wate Cirltk Banel Reference Wata Cir Auto Bin iti hres Auta Panel iti hres Edit SNP Marker The Edit SNP Marker command opens the Edit SNP Marker dialog box and allows you to edit the marker name and bin information for the selected marker To enable this feature e Select the menu item when a SNP genotyping marker is selected in the Panel Manager Plot tab or e Right click on a SNP genotyping marker or bin in the Panel Manager Plot tab 4 12 GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands You can quickly edit SNP genotyping markers and bins by changing the data in the Panel Manager Table tab for a selected SNP genotyping Panel Delete SNP Marker The Delete SNP Marker command deletes the SNP genotyping marker and associated bins selected in the Panel Manager Plot tab To enable this feature e Select the menu item when a SNP genotyping marker or bin is selected in the Panel Manager Plot tab or e Right click on a SNP genotyping marker in the Panel Manager Plot tab Add Bin The Add Bin command creates a new bin associated with a panel as follows For microsatellite data 1 Select a marker in the Panel Manager navigatio
261. ng data generated with AmpF STR kits using forensic simulated and single source DNA samples and electrophoresed on ABI PRISM platforms Our findings demonstrate that GeneMapper D Software v3 1 with its default settings and panels and bin sets is valid for forensic paternity and databasing analyses This software accurately detects genotypes and performs quality checks on data analyzed with GeneMapper ID software However during the verification testing the process component based quality value PQV flags for spectral pull up peaks associated with off scale peaks can be inaccurate Further our findings demonstrate that the genotypes between GeneScan and Genotyper software to GeneMapper JD software are concordant Laboratories currently using GeneScan 3 1 2 for the Macintosh OS can maintain current analysis parameter settings when adopting GeneMapper JD software in the Classic mode Applied Biosystems verification testing demonstrates that human identification laboratories can successfully adopt GeneMapper ID Software v 3 1 GeneMapper ID Software v3 1 User Guide I 39 Appendix GeneMapper ID Software Verification References l 40 Applied Biosystems 2001 ABI PRISM AmpFSTR Identifiler PCR Amplification Kit User s Manual Applied Biosystems 2002 ABI PRISM GeneScan Analysis Software for the Windows NT Operating Systems Overview of the Analysis Parameters and Size Caller Boland C E Leibelt C S Daoudi
262. nly These two flags are independent of one another Therefore the thresholds are set independently The software calculates a numeric value from 0 to 1 to represent the sizing quality and genotype quality based on certain criteria and or PQVs You can then determine a numeric low quality range and pass range For example if you use the default threshold settings shown in the figure below any results over 0 75 pass and any results below 0 25 are of low quality Any results between 0 26 and 0 74 are marked to check Use the Quality Flags tab to set the importance level of the PQVs available for HID and SNP genotyping analysis The individual PQVs on this page are used in the calculation to determine the overall genotype quality This tab allows you to determine how much weight is applied to each PQV in this calculation GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab Use the options under PQV Thresholds to set Pass Check Low Quality thresholds for the Sizing Quality SQ parameter for the Samples view and the Genotype Quality GQ parameter for the Genotypes view Anything that is not labeled Pass or Low Quality will be labeled as check A Note Applied Biosystems recommends that you use the default settings for AmpF STR data analysis Options Data entry restrictions for the PQV thresholds ensure that the lower limit of the Pass range is always greater than the upper limit of the Low Quality range The
263. no instance of the type specified exists the Genotypes table will be displayed blank Return to the Table Manager and reset the filter GeneMapper ID Software v3 1 User Guide 5 37 Chapter 5 Using GeneMapper Manager The following information describes how to use the filter settings for the Genotypes Column Settings The default Show All Records can be changed to one of the following Table 5 11 Genotypes view filter settings Column Name Filter Settings Sample File Show Records Containing Sample Name Sample ID P Run Name Show All Fecords Panel i Marker Show All Records a Show Records Containing Size show Records Containing Height Note This setting allows specification by a Peak Area string Input this information into the Content Mutation column AE Comment Integration Comments User Defined Columns Dye Blue Green Orange Red or Yellow Show All Records Show All Records Note This setting can be changed to display either all colors Show All Records or the choice of display of an individual color SNP genotyping data cannot be filtered by this selection 5 38 GeneMapper ID Software v3 1 User Guide Table 5 11 Section 5 3 Table Settings Tab Genotypes view filter settings continued Column Name Filter Settings Off scale Sharp Peak M One Basepair Allele M Single Peak Artifact M Split Peak M Out of Bin Allele Peak Height R
264. nts Show All Records 6 p Sample Type Show All Records N A 7 p Specimen Category Show All Records N A 8 m Analysis Method Show All Records Plot window a sample or results row and then either select Analysis gt Display Plots press Ctrl L Alternative Click the icon m Plot Window Samples Plot lol xj File Edit View Tools Alleles Help Plot Setting Ho Genotyping z Panes fa z EG Ei Bi al WE E ed EE F E AES Sample Name Pand sgo sQ D851179 D21511 D75820 CSFIPO 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 200 0 a Laj faa pho io l Samples entifitervi C D3 1358 THO D135317 D165539 D251338 400 110 120 130 1440 1450 4160 170 180 190 200 20 220 230 240 250 260 270 280 290 300 310 320 330 340 350 Ee es ee er eS ce oe a a or Se Se SS a a ST Sen 400 o D19S433 DIsS51 100 110 120 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 sampie denier vt E An D55818 GA 100 110 420 130 140 150 160 170 180 130 200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 i 0 e Bd hais ba e iE Lal EES ee 183 35 Y 191 GeneMapper ID Software v3 1 User Guide Chapter 3 Using the Project Window Project Window Toolbar ee
265. nts and descriptions Element Description OK Accepts the pending changes and closes the dialog box The following criteria apply e The edited bin must have a name unique in its marker e The edited bin must have a location and offsets that do not overlap any other bins in the marker An alert message will be displayed if these criteria are not met Cancel Closes the dialog box without accepting the pending changes New Offset Checking this box sets a left and right offset Default default to be used for subsequent creation of bins checkbox Mutant Bin Use this feature to distinguish microvariant bins from known allele bins Bin color is red New Bin Set New Bin Set a X amp Please enter name of new Bin Set The New Bin Set command opens the New Bin Set dialog box allows you to create a new bin set for the selected kit Clicking OK checks the name for uniqueness if the name is not unique an alert is shown and the dialog box closes The new bin set name then displays at the top of the Panel Manager window in the Bin Set pull down menu Note Bin sets are always created at the kit level Note SNaPshot kits can only contain one bin set GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands Delete Bin Set F Delete Bin Set The Delete Bin Set command deletes the bin set currently listed for a kit in the Bin Set drop down menu at the top of the Panel
266. o Applied Biosystems To contact Applied Biosystems technical service and support see How to Obtain Support on page xi in the Preface IMPORTANT Installation privileges are available only if you have returned your registration card To register GeneMapper ID software l Select Start gt Programs gt Applied Biosystems gt GeneMapper gt GeneMapperID v3 1 or double click the GeneMapperID v3 1 desktop shortcut E GeneMapper ID 3 1 Product Registration Your Hame Organization Registration Code Quit 2 Register the software a In the Product Registration dialog box enter your name organization and registration code on the registration card b Click OK GeneMapper D software then verifies the code Note Keep your registration code in a place where you can easily retrieve it If you need to reinstall the software at any time you will be prompted for the registration code again GeneMapper ID Software v3 1 User Guide Logging Onto and Logging Out of the GeneMapper ID Software Logging Onto and Logging Out of the GeneMapper ID Software Introduction GeneMapper D Software Access Rules Logging On This section provides information on logging on to and logging out of the GeneMapper JD software Follow these rules to access the GeneMapper ID software e The first time you launch the GeneMapper ID software application after installation you must register the soft
267. o your Macintosh computer s CD ROM drive An icon displays for the CD ROM on the right hand side of the screen 2 Double click the CD ROM icon A CD ROM window displays containing files and folders 3 Double click the CONVFOLD folder Inside this folder are two files CONVPROG HOX and README TXT which contain the installation instructions 4 Copy the CONVPROG HOQX file to your local hard drive by click drag the file over to the local hard drive icon and releasing 5 To decompress the CONVPROG HQX file drag and drop the file on the Stuffit Expander icon Boi Note You can download a free version of Stuffit Expander from http www stuffit com expander GeneMapper ID Software v3 1 User Guide Converting Macintosh Sample Files Note Decompressing the CONVPROG HQxX file creates a Conversion Programs folder on the local hard drive This folder contains the conversion programs and the SimpleText file About Conversion Programs This file is a seven page document that describes in detail how to use the conversion programs why they are necessary solutions to common problems and possible alternative programs GeneMapper ID Software v3 1 User Guide F 3 Appendix E Converting Macintosh Sample Files Converting To convert Macintosh computer sample files for use on a Macintosh Computer running Microsoft Windows operating system Sample F iles to 1 Double click the H gt icon to start the program Micros
268. of the matched size standard in the tabular data Partial Sizes the software reports peaks within this user defined size range in the tabular data Note To ensure successful sizing the analysis range should contain all size standard fragments includes in the sizing range For AmpF STR data analysis follow the below chart for recommended sizing ranges Table 5 6 Recommended Sizing Ranges for AmpF STR Data Analysis Kit Sizing Range e AmpF STR Profiler Plus kit 75 400 e AmpF STR COfiler kit e AmpF STR Profiler kit e AmpF STR Blue kit e AmpF STR Green kit e AmpF STR SGM Plus kit e AmpF STR Identifiler kit 75 450 e AmpF STR SEfiler kit Peak Amplitude Thresholds The values you enter for each color represent the minimum peak height in relative fluorescent units that the software will detect and size The software ignores any peak heights below this value For all dyes the default is 50 GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab For more information and a detailed description of all other parameters refer to Appendix B Software Genotyping Algorithms Note Applied Biosystems recommends that you use the default settings for all other analysis parameters for AmpF STR data analysis Classic Peak Detection Algorithm This algorithm is similar to the GeneScan software on the Macintosh operating system m
269. oft Windows Files Sample File Aac to Wiin The following dialog box opens U USS Enterprise am bate ried a B ea Modules af 20 02 GT Runs Today C Fun Folder 04 30 03 07 20 AM 5 21 03 C Fun Folder 05 07 03 03 32 PM 5 72 03 E Ekun Folder 05 15 03 10 19 AM 5 15 03 Where are the files to convert to Windows format Note On Macintosh computers running operating system 8 0 or less this dialog box has a different appearance For more information refer to the SimpleText file About Conversion Programs mentioned above in Installing Conversion Programs 2 Using the triangle shaped icons to the left of the folder names navigate to the folder that contains the fragment analysis sample files you want to convert 3 Select the folder 4 Click Choose If there are no problems the program performs the task and quits automatically When you open the folder the sample files have the file extension fsa Note To convert sample files created on a computer running the Microsoft Windows operating system for use on a Macintosh computer follow steps 1 3 above in Step 1 double click the Sample File Win to Mac icon E 4 GeneMapper ID Software v3 1 User Guide Transferring Data and Backing Up Data This appendix covers Transferring Data 0004 Backing Up Data and Cleaning the Database GeneMapper ID Software v3 1 User Guide Appendix F Transferring Data and Backing
270. on The large peak is excluded from the candidate list by a filter that has identified it as being atypical with respect to the others Bars indicate size standard determined by algorithm Bars indicate peaks found JOO herder ces deren e o pa E EEA EA E E seese eilei i m cea flourescence 800 1600 scan line Binning Algorithm This algorithm estimates the bin centers using all found alleles To determine true bin positions the algorithm uses an iterative clustering technique and then reports a quality value proportional to the binning noise To enhance flexibility bins may also be edited manually Note When you create bins manually all alleles regardless of their quality values are treated as reference alleles as long as the alleles reside within the created bins boundaries Allele Calling Final allele calls are based on a consensus between several different Algorithm allele calling algorithms Each caller has a different design philosophy such that it excels in a particular data regime A variety of allele calling algorithms are used Allele calling algorithms involve envelope detection optimization of parametric models and rule based systems GeneMapper ID Software v3 1 User Guide B 3 Appendix B Software Genotyping Algorithms Example Output of Different Allele Calling Algorithms The following is an example of three different allele calling algorithms for 16 samples User annotations are
271. on in base pairs for any selected bin are shown under the X axis to the left The Bin Sets drop down menu located just above the Bin view pane selects the bin set that is applied to the marker Marker Allele Size Range ASR Reference data for bins includes the marker ASR This is the size range of bins in a marker to allow multiplexing of markers in a sample ASR boundaries can be edited in the marker table or in the Bins view by adjusting the ASR handles on the right and left hand sides of the screen The bins in a marker must not extend beyond the marker ASR When bins are created and viewed in the Bins view they are restricted so that their range does not extend beyond the marker ASR The Show Alleles command may show new allele calls for a marker GeneMapper ID Software v3 1 User Guide 4 41 Chapter 4 Using Panel Manager outside that marker s ASR but bins cannot be created or extended to include alleles outside the ASR To create bins for alleles that fall outside the ASR edit that marker s ASR click Apply and add a new bin Bins must not overlap in a microsatellite marker When bins are created and edited their locations and offsets are restricted to avoid overlap Import and export commands for panels and bin sets are enabled in the table views of the Panel Manager Imported bin set reference data includes the bin boundaries but not sample reference data the s 4 42 GeneMapper ID Software v3 1 User Gu
272. only used when creating panels manually not when using the Auto Panel feature The panel is enabled when a SNP genotyping panel is selected and reference data has already been added at the kit level e Microsatellite data Use the Panel Reference Data command to remove reference samples from the selected microsatellite panel Auto Bin F Auto Bin E e xj Minimum quality value 0 4 Allele Naming Scheme Number 1 2 3 4 2 3 Letter 4 6 0 3 gt Rounded basepair 100 200 300 Auto Bin Options i Auto Bin clear existing bins C Incremental Auto Bin keen existing bins oox caeai The Auto Bin command opens the Auto Bin dialog box to automatically create bins for a microsatellite panel This feature is enabled when a microsatellite panel is selected and reference data has been selected Refer to the Microsatellite Tutorial for more information 4 18 GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands Auto Panel Auto Panel ee E X Hee ot Save Existing Panels Allele Names Meu fo Yellow Jef Mcreen fa E Red qT Minimum multiplex E Maximum multiplex j0 Panel Hames base Panel name Marker Overlap wn Maximum overlap No overlap Mumber of basepairs between markers 5 OK Cancel The Auto Panel command opens the Auto Panel dialog box to automatically c
273. ontains a single ROX fluorophore The following table lists the lengths of the 21 fragments that make up the GeneScan 400HD Size Standard Fragment Fragment Fragment Length Length Length 50 180 290 60 190 300 90 200 320 100 220 340 120 240 360 150 260 380 160 280 400 Although the GeneScan 400HD Size Standard is made of double stranded DNA fragments only one of the strands 1s labeled Consequently even if the two strands migrate at different rates under denaturing conditions you will not need to worry about peak splitting GeneMapper ID Software v3 1 User Guide D 5 Appendix D GeneScan Size Standards Electrophero gram of GeneScan 400HD gt Size Match Editor The following screen shows an electropherogram of GeneScan 400HD Edit View 3094 D 6 Size Match Quality 1 0 240 260 280 290 300 150 160 180 190 200 220 3610 3868 4126 4584 a SS SB 5674 5932 6190 64de 6706 6964 7222 7480 a ea Lao GeneMapper ID Software v3 1 User Guide GeneScan 500 and GeneScan 500 250 Size Standards GeneScan 500 and GeneScan 500 250 Size Standards About These Size The GeneScan 500 the GeneScan 500 LIZ the Standards GeneScan 500 250 and the GeneScan 500 250 LIZ Size Standards are useful for sizing fragments between 35 and 500 base pairs The native fragments are uniformly spaced to pro
274. op a set of criteria to evaluate the performance of this software for the human identification communities The verification was designed to evaluate four specific categories e Peak detection and genotyping e Algorithm testing e Workflow e Data handling These categories cover 24 test parameters that evaluate the software s robustness performance and features The test parameters were performed using forensic simulated samples amplified with the AmpF STR PCR Amplification kits and electrophoresed on defined ABI PRISM instrument platforms These test parameters evaluate for example peak detection and the non concordance of alleles out of bin alleles peak height ratios low peak heights the ability to process samples per color per sample and per marker and import and export functionality GeneMapper ID Software v3 1 was specifically designed as a complete integrated software package for forensic paternity and databasing laboratories performing STR analysis Currently human identification laboratories worldwide use ABI PRISM GeneScan Software and ABI PRISM Genotyper Software as the standard for software analysis GeneMapper JD software combines the functionality of these two software programs into one integrated software package Among other things this software analyzes the raw data collected from ABI PRISM instrument platforms and automatically identifies peaks quantifies signal intensity sizes each DNA fragment
275. oring Sample Files Projects and Second Reads The following workflow applies only when the computers running GeneMapper D software are connected to your network 1 Verify that all copies of the GeneMapper JD software in the lab have exactly the same analysis method size standard definition panels binset and matrix if applicable imported into the GeneMapper Manager a From the GeneMapper Manager export the master copy of each of the files to a designated location on a server b For each computer using GeneMapper JD software use the GeneMapper Manager to import the files from the server c Keep the master copy of the files on the server as a backup 2 Store all sample files on a server that can be accessed by all computers with GeneMapper ID software Do not move these sample files or download them to personal computers Note The GeneMapper project requires an unaltered link to the sample files If these sample files are moved the link 1s broken and some functionality within the project such as accessing the raw data viewing certain plots and re analyzing samples is lost 3 For the initial analysis of the data by the first or lead analyst import the sample files into a new project perform analysis edit the alleles and save the project to a designated location on the server 4 For the second analysis of the data Using Same Computer Then Proceed to Step Yes 5 and skip step 6 No
276. orithms B 2 H HID analysis considerations 7 2 HID analysis options 6 3 GeneMapper ID Software v3 1 User Guide Import Project window 5 7 IMPORTANTS description x Info tab 3 6 Italic text when to use 1x L Least Square method B 29 Local Southern method B 32 logging out of GeneMapper 2 10 M Macintosh sample files converting E 2 marker allele size range ASR 4 41 Marker Table view 4 36 matrices about 5 54 causes for bad 5 64 error messages 5 64 limitations 5 55 where to store 5 56 5 64 matrix assigning to a sample 5 55 assigning to sample 5 63 choosing a data point range 5 58 5 59 creating a GeneMapper 5 60 definition 5 54 eliminating primer peaks 5 59 example 5 62 generating anew 5 60 process of creating anew 5 57 raw data display 5 59 steps to create 5 58 using different computers for data collection and analysis 5 63 using with data collection 5 63 viewing the raw data 5 58 when to assign to a sample 5 55 when to create 5 56 matrix files about 5 54 to 5 56 bad matrix files causes 5 64 choosing a data point range 5 58 to 5 59 GeneMapper ID Software v3 1 User Guide new matrix file generating 5 60 to 5 62 process of creating new file 5 57 Menu commands conventions for describing 1x MSDSs obtaining xi N Notes description x O off ladder alleles 7 3 Open Project dialog box C 28 Options dialog box 6 2 C 18 options setting 6 3 P Panel and Bin text file
277. ory Provided HID Genotyping 2003 06 12 14 45 4 grid HD Last Used Plot 2003 06 1214 534 gmid Mew Open Save As Import Export Delete Figure 5 22 Plot Settings Tab of the GeneMapper Manager ltem Description Name column Plot setting name Last Saved Date Time stamp showing when the plot setting column was last saved Owner column User name of the person who created the plot setting Description A description of the plot setting column GeneMapper ID Software v3 1 User Guide 5 41 Chapter 5 Using GeneMapper Manager Default Profiles 5 42 Table 5 12 Description of elements in the Plot Settings Tab Item Description New button Opens the Plot Settings editor Always enabled Open button Save As button Import button xml file extension Opens the editor for a selected project Enabled when a single plot setting is selected Displays the Save As dialog box Enabled when one or more settings are selected Displays a dialog box for Importing plot settings Always enabled Export button xml file extension Displays a dialog box for Exporting selected plot setting Enabled when one or more plot settings are selected Delete button Deletes the selected plot setting s Done button Closes the GeneMapper Manager Applied Biosystems provides five default plot settings for AmpF STR analysis e AmpF STR Genot
278. ou to choose where to save the Dialog Box exported file The file types supported include e Tab delimited text txt e Comma separated values csv Note Export operates on the currently displayed tab For example if the Samples tab is displayed Export would export the tab delimited data shown in the Samples table Exported data includes column headers 1 2 Export Table Look in __ GeneMapper Export File 45 3 Tab delimited text tot ze 8 Recent Perce Example Data Desktop GM3 1 Database Defaults rn installer ee JRE My Documents Panels Plot Settings Size Standards _ Table Settings GeneMapper Log Panellmportlog 4 Fiene EEA Tee Files of type frav and Comma delimited Text txt cav ha Cancel 7 GeneMapper ID Software v3 1 User Guide C 31 Appendix C Project Window Software Interface C 32 Export table callouts Item Name Description 1 Toolbar Click these icons to Move display up one level in main pane Move display to Home level in main pane This is usually Profiles lt user gt Create a new folder at the present ag directory level ee Present a list of the contents of the B n selected folder Present details of the selected folder 2 Drop down Select drive letter and or folder directory menu 3 Folder file The contents of the folder selected in the display pane toolbar are displayed he
279. ou to create edit and import Panel Manager kits panels markers loci bins and bin sets It features the Window _ following e A tool for creating kit folders e A table for specifying panels e A table for defining loci within a panel e A graphical editor for creating and editing bins All information imported entered or created is stored in the database This information includes kits panels markers loc1 bins and bin sets e Clicking Apply after making changes to a panel sends all changes to the database but keeps the window open for further edits e Clicking OK sends all changes to the database and closes the Panel Manager window The changes in panel data are then reflected in the Project and Plot windows after the samples are re analyzed When to Click the e After deleting a kit panel locus or bin set Apply Button e After making a number of edits e After creating a bin set e After making any change you want to save GeneMapper ID Software v3 1 User Guide 4 3 Chapter 4 Using Panel Manager 4 4 Displaying the Panel Manager Window Hierarchy of Panel Data To display the Panel Manager window either e Click W Panel Manager from the Project window toolbar or e Select Tools gt Panel Manager in the Project window r7 CA Panel Manager File Edit Bins View E E Panel Manager Farel Harte SE aF AmpFLSTR Panes iE Beo froe ABlue_v1 2 cr vd none Lsreen_ l1 3 Profiler none H E Frotil
280. ovides information about the ranges of possible values Table 5 5 Value ranges for tetranucleotide Parameter Min Max salle aloes Cut off value 0 0 1 0 0 0 MinusA ratio 0 0 1 0 0 0 MinusA distance 0 0 0 0 Stutter ratio 0 0 1 0 0 0 Stutter distance 0 0 from 3 25 to 4 75 bp Amelogenin cut off 0 0 1 0 0 25 Range Filter The Range Filter allows you to remove labels from peaks within a specific size range for each color 5 18 GeneMapper ID Software v3 1 User Guide Section 5 2 Analysis Methods Tab Peak Detector Use the Peak Detector tab to select the algorithm to use for detecting Tab _ peaks e Advanced e Classic The Peak Detector tab with Advanced and Classic peak detection algorithm settings are described in the following pages See Appendix B Software Genotyping Algorithms for additional information on Advanced and Classic peak detection algorithms Advanced Peak Detection Algorithm This algorithm is similar to the method of analyzing size standards and performing peak detection that is used in the ABI PRISM GeneScan software on the Windows NT operating system Information on the individual parameters can be found in Appendix B Software Genotyping Algorithms Information about ranges and peak amplitude thresholds is as follows Analysis Method Editor HID Peak Detection Algorithm advanced z Ranges Feak Detection Analysis S
281. ow by performing standard actions like undo delete select and by enabling access to settings for Preferences sale analysis View Tools Delete from Project Select All Cirlt A Fill Down Ctrl D Find Ctrl F Sort Ctrl G Item Description Enabling Delete From Deletes the selected sample s Enabled when a Project from the Project and the database Project sample is and displays the following alert message Deleting the selected sample s will delete both samples and results from project This action cannot be undone OK Cancel selected Note Each use of the command removes a single run or sample If you want to remove all samples and the associated folder use the Select All command before using the Delete from Project command This action does not delete the actual sample file or sample data GeneMapper ID Software v3 1 User Guide Project Window Menus Item Description Enabling Select All Ctrl A Selects all samples in the active view Always enabled Fill Down Ctrl D Fills the selected column with the contents of the first cell This command is applicable only to Analysis Method Panel Size Standard and Matrix columns Enabled when a column is selected or multiple cells are selected Find Ctrl F Sort Ctrl G Displays the Find dialog box Displays the Sort dialog box Enabled when the table contains data Enable
282. p Data Cleaning the GeneMapper Database The GeneMapper database stores all projects not including sample files size standard definitions analysis methods plot and table settings panels bins and matricies Therefore it 1s recommended that you back these components up in case of a computer failure or crash In order to do this you must export each component individually through the GeneMapper Manager and the Panel Manager By exporting the components they are not deleted from the database In order to delete the items you must select them within the GeneMapper Manager or Panel Manager and select delete The GeneMapper software may potentially perform slower as the GeneMapper database fills ups Therefore it is recommended that you back up all of your projects and delete them from the database periodically Keep in mind that when you export a project the sample files are not linked Therefore if you need to re import the project at a later date you will need to follow the instructions above in the transferring data section GeneMapper ID Software v3 1 User Guide F 7 Appendix F Transferring Data and Backing Up Data F 8 GeneMapper ID Software v3 1 User Guide Sample Sheets and Plate Records The appendix covers Information from the Data Collection Sample Sheet Or Plate RECON 6 255 in ed Sra eu heed ey a E era Ader A G 2 GeneMapper ID Software v3 1 User Guide G 1 Appendix G Sample Sheets and Plate Record
283. peak is not off scale and should be flagged green the spectral pull up PQV flag is green this is true spectral pull up and should be flagged yellow and the broad peak PQV flag is yellow this peak is not broad and should be flagged green As mentioned these PQV flag colors for off scale spectral and broad peak should be reversed for the pull up peak but the PQV for the true off scale peak is accurate The out of bin PQV flag for all tested alleles accurately reflect the data GeneMapper ID Software v3 1 User Guide l 27 Appendix I 28 GeneMapper ID Software Verification ile Edit View Tools Alleles Help Plot Setting HD Genotyping I EI Paes I E A E al dF el FA E a Figure l 17 Example of an off scale allele in the Advanced mode Note the off scale display the pink bar represented in the same size position in the panels above and below the off scale allele The pink bar does not accurately reflect off scale data in the top and bottom panels but only reflects an off scale allele in this sample at this position la x File Edit View Tools Alleles Help E Pones fF T_T li Ea Plot Setting G5_3100 r fe LP 2a Sample Name Pand SQ0 SQ Spectral pull up in the THO1 size range ls AN A AA E A en A E A AE EE E Eni H i E i Bl m i Figure I 18 For this Identifiler example a pink bar is displ
284. pen the bin edit options and select Edit Bin The Edit Bin dialog box opens allowing you to edit bin information To quickly resize a bin select a bin and adjust the right and left handles to the desired size range To relocate or move the bin mouse action only 1 Select Binning mode 2 Select the bin to be moved 3 Click and drag the center of the bin to a new location GeneMapper ID Software v3 1 User Guide 8 27 Chapter 8 Using Plot Windows Samples and Genotypes Alleles menu items and descriptions continued Item Description Delete Bin Delete Allows you to delete a marker bin as follows 1 2 3 Select Binning mode Select the bin to be deleted Select the Delete Bin menu item or right click the selected peak to open the bin edit options and then select Delete Bin The bin will then be deleted Move Allele Using the mouse you can move an allele label to a new peak To move an allele 1 Left click a label to select it 2 Hold down the Ctrl key and drag the label to the new peak The old label will be marked removed The new label will be marked added Help Menu The Help menu provides access to this user guide in PDF and to the About GeneMapper ID software window GeneMapper Help F1 About Genehlapper 49 Help menu items and descriptions Item Description GeneMapper Help F1 Opens the GeneMapper ID User Guide PDF file us
285. ples Databasing Identifiler_v1 AAA Y p7 E E aa E S a E i 92 Samples Databasing Identifiler_v1 TPOX Y l8 9 a E E a E E A a a 93 Samples Databasing Identifiler_ 1 D18551 Y 15 18 Ni E E E E E a E i 94 Samples Databasing Identifiler_v1 AMEL R X G a a E B i E B ia 95 Samples Databasing Identifiler_ 1 D55818 R 11 13 o E A E m A iai a B 4 gt 96 Samples Databasing Identifiler_v1 FGA R 24 26 E E E E fi E E g Analysis Completed E Figure l 19 Genotypes table of the CSF1PO locus of a 1 to1 mixture displaying two yellow PQV flags Peak Height Ratio and Allele Number giving an overall genotype quality value of red GeneMapper ID Software v3 1 User Guide I 29 Appendix GeneMapper ID Software Verification As illustrated in Figures I 19 the CSF1PO locus is displaying a yellow flag for both PHR and AN By default the setting for PHR is 70 and the maximum number of alleles expected is 2 Figure I 20 displays the electropherogram of the CSF1PO locus selected in Figure I 19 In Figure I 20 the PHR is below the default setting and the maximum number of expected alleles is greater than the default setting 2 The software has appropriately flagged the CSF1PO locus for the PHR and AN PQV F Genotypes Plot File Edit View Tools Alleles Help Plot Setting Ho Genotyping x Panes fa z PET EEr
286. ples must be analyzed and the project saved e The sample files to be compared must have the same name Note If the sample file names are different due to laboratory protocols or automatic naming conventions you can still perform this procedure by typing a shared name for two or more samples in the UDI column You can simply highlight the cell in the UD1 column and type in a common name for the samples to be compared This information can also be read from a sample sheet If you type a pipe bar after the panel name in the comment field Profiler Plus vl samplel the information after the pipe bar will be displayed in the UD1 column To check concordance for shared markers 1 Ifthe genotypes table is not already displayed click on the genotypes tab in the main project window or select View gt Genotypes 2 Select Analysis gt Non concordant Sample to Top Any rows that are non concordant will be brought to the top and highlighted 3 To view the plots for all non concordant markers keep the rows highlighted and select Analysis gt Display Plots GeneMapper ID Software v3 1 User Guide Using Plot Windows samples and Genotypes This chapter describes how to examine and interpret electropherograms within the GeneMapper ID Software Version 3 1 Samples and Genotypes plot windows and explains the various electropherogram settings and interactions in the plot window menu items This chapter covers About th
287. plet tsa Clear Ada Add amp Analyze Cancell Element Description Files pane Contains the directory structure configured for the computer running GeneMapper D software The navigation pane provides a tree viewer that works like the MS Windows Explorer interface You can use Shift click for continuous multiple selection and Ctrl click for discontinuous multiple selections within a folder Samples To Add pane Contains the folders and files that the system will add This field also uses the tree viewer interface You can use Shift click and Ctrl click as described above Add To List button Adds the selected items from the files pane to the Sample to Add field Options button Opens the Options window Clear button Clears the selected files from the Samples To Add pane Add button Adds the Samples To Add list into the Project window Cancel button Closes the Add Samples to Project dialog box GeneMapper ID Software v3 1 User Guide C 41 Appendix C Project Window Software Interface GM Database Tab The GM GeneMapper JD Database tab enables you to choose sample files that have previously been added into a GeneMapper ID project This database uses a search interface to extract samples that match your criteria Criteria Selector list Samples To Add pane Search Criteria Table Add Samples to Project Edit View Files GM Dat
288. ponent Based Quality Values A 3 GeneMapper ID Software v3 1 User Guide A 1 Appendix A Process Quality Values Overview of Process Component Based Quality Values PQV What are Process Process Quality Values PQVs values reported by data analysis aid Quality Values in finding and fixing problems in sample preparation and analysis These values are the end results reported by the PQV system The PQV Process The PQV process includes the following e At the end of the process quality values also known as PQV or Process Component Based Quality Values are assigned to the size calling process and the allele calling process e On the Samples page and Genotypes page the columns presenting PQV result data other than SQ and GQ results display the following samples after analysis Pass Ii green square symbol when no problem exists Check A yellow triangle symbol when there are problematic components such as missing size standards or missing matrices Low Quality red octagon symbol when the result falls below the defined threshold What are PQVs PQVs reported by data analysis are an aid to finding and fixing problems in sample preparation and analysis These values are the end results reported by the PQV system Editing PQVs_ When you edit PQVs you must create an Analysis Setting and select the application mode to control which PQVs are used and available to you for editing A 2 GeneMapper I
289. profile entered at a time per panel Note Ifa ladder injection is of low quality you can change the sample type from Allelic Ladder to Sample This will remove the low quality injection from consideration in calculating the allelic bins 2 Inthe Analysis Method column select a method GeneMapper ID 3 1 Untithed gmid Is Logged In File Edit Analysis View Tools Help e coe Sy al GA aE E Tae s El Project samples Genotypes H E Casework Specimen Cater Analysis Method oi ia noexport fro expet no export oo j CS a Use the drop down list to select the appropriate method Note You have the option to define a new analysis method from this drop down list b Click the column header cell to highlight the entire column or highlight a portion of the column that you want to apply the same setting to then select Edit gt Fill Down GeneMapper ID Software v3 1 User Guide 7 7 Chapter 7 Analyzing and Interpreting Data 3 In the Panel column select a panel a Highlight a cell within a sample row by clicking it in the panel column A Select a Panel window will appear b Expand the AmpFLSTR Panels v1 folder Green_ _ v1 Protiler_ v1 Protiler_Plus_ 1 Cofier Ssh Flus _w Identifier v1 SEtiler_ v1 Profiler Plus CODIS 1 Copier CODIS_ 1 Identifier CODIS_ 1 c Double click the appropriate panel to populate the cell within the panel column d Click the colu
290. project was last saved User name of the person who created the project Number of Samples column Number of samples contained in the project Rename button Opens a Rename dialog box for renaming the project Enabled when a project is selected Save As button Displays the Save Project As dialog box Enabled when a project is selected Import button Displays a dialog box for Importing projects Export button Note This exports projects only not the sample files Displays a dialog box for Exporting selected projects Enabled when projects are selected Note Projects can be exported as a group by holding down the shift key and selecting multiple projects This feature works with one or more selected projects Delete button Deletes the selected projects Enabled when a project is selected Note This feature works with one or more selected projects Done button Closes the GeneMapper Manager GeneMapper ID Software v3 1 User Guide Section 5 1 Projects Tab Export Project Use the Export Project window to export a project out of the Window GeneMapper JD database into a specified location See an explanation of each area on this window in Using the Export Project Window on page 5 6 Note The only method of backing up your projects is to export them _h 2 F Export Project Look in GeneMapper 3 Examples JRE 4 5 File na
291. ptions Item Description Smoothing Used to help reduce the number of false peaks option buttons detected by the GeneMapper ID software You have the following options Select To None apply no smoothing Select this option if the data has very sharp narrow peaks of interest Light provide the best results for typical data Heavy apply to data from slower runs that has very broad peaks or to avoid the detection of sharp edges Choosing this option might reduce peak size or eliminate narrow peaks Baseline Window Used to automatically adjust the baselines of all text field detected dye colors to the same level for a better comparison of relative signal intensity Baselining Option The Baselining option controls the scope of the baseliner Use this option to set the size Beta of the Baseline Window GeneMapper D software computes a baseline for the electropherogram of each dye independently How the Baselining Option Works A baseline comprises a value at each data point i The baseline value at each data point i is the lowest electropherogram value in a window whose width Beta is set using the Baselining option and centered at each data point i GeneMapper ID Software v3 1 User Guide B 13 Appendix B Software Genotyping Algorithms More accurately the baseline computed in this manner is intermediate The real baseline value at each data point 7 is the highest interm
292. r ID Software v3 1 User Guide 4 39 Chapter 4 Using Panel Manager Description of the Bins View Reference Data Bins and Allele Calls Reference data includes descriptions of bins name location and boundaries and the sample allele calls used to construct the bins In the Bins view the vertical bars are bins that are originally associated with the markers imported as reference data The symbols shown in the figure above marked in red on the monitor are from the reference alleles used to build the bin set indicated in the drop down menu at the top of the window The red s are shown when those alleles are used in the autobinning process or in the manual process of bin creation Panel Manager File Edit Bins View a X E E ET 4 Bin Set ampFLSTR_Bins_v1 al Eana loo e 4 JSNP MICROSATELLITE_Y E5 AmpFLSTR_Panels_v CBlue_v1 CI Green v1 Cf Profiler _v1 El E5 Profiler _Plus_v1 fe 2381358 m D21511 D18551 gt D55813 a D13S317 D75820 bea b EJ Reference Samples 4 4 7 3 9 10 11 12 13 14 15 16 Le 18 19 20 1 0 0 8 0 7 0 6 05 0 4 0 3 0 2 0 1 0 0 Ts Tie a D851179 125 129 133 137 141 145 149 153 157 161 165 169 173 177 181 xX 126 81 Y 0 50 OK Cancel Apply 4 40 GeneMapper ID Software v3 1 User Guide Bins View Microsatellites Only If you analyze data using a bin set like that shown above you will see the s as shown above T
293. r than a certain value The default of this value is 1 5 basepair This flag is used for HID SNP and Microsatellite all markers Not editable SP Split Peak e A flag Pass Ii or Check A is automatically displayed when a split peak is found by the GeneMapper ID software A split peak is defined as overlapping peaks that are lt 25 basepairs apart the horizontal distance from two peak apexes Not editable GeneMapper ID Software v3 1 User Guide A 9 Appendix A Process Quality Values The PQV flags on the Genotypes page continued Column Description OS Offscale e A flag Pass Ii or Check A is automatically displayed when there are offscale peaks present within the marker size range e This flag is used for all applications e Not editable CC Control Concordance e A flag Pass Ii or Check A is automatically displayed when the designated control sample does not exactly match the defined alleles for this marker in the panel being scored e This feature serves as an internal control for quality assurance and allows you to see deviations in your controls e This flag is used for all applications e We recommend that you run the control sample at least once for every panel e Not editable OVL Overlap e A flag Pass oor Check Ad is automatically displayed when a peak in the overlapped region is called twice It is possible to have two allele size ranges that ov
294. r Classic and Advanced Modes Overview There are four size calling methods that you can use with the GeneMapper D software v3 1 They are as follows e Least Square e Cubic Spline Interpolation e Local Southern e Global Southern Least Square Both Least Squares methods 2nd Order and 3rd Order use Method regression analysis to build a best fit size calling curve This curve compensates for any fragments that may run anomalously As a result this method normally results in the least amount of deviation for all the fragments including the size standards and the samples Depending on whether you choose the 2nd or 3rd Order Least Squares Method in the Analysis Parameters dialog box the resulting size curve 1s either a quadratic or a cubic function The software uses the known standard fragments and the associated data points to produce a sizing curve based on Multiple Linear Regression Advantages In Figures B 10 and Figure B 11 on page B 30 you can see that in nearly all instances the mobility of an individual DNA fragment is coincident with the best curve fit of the entire data set Stated differently the mobility of most DNA fragments 1s strictly length dependent This method automatically compensates for fragments that run anomalously GeneMapper JD software v3 1 calculates a best fit least squares curve for all samples regardless of the size calling method you choose The curve is black in the Standard Sizing Curve window
295. r Not Analyzed Show All Rec wY Show All Recards Analyzed Mot Analyzed Sample Name Sample ID Comments Analysis Method Panel Size Standard Matrix Run Name Instrument Type Instrument ID Run Date amp Time User Defined Columns Show Records Containing Show All Recards Show All Records Show Records Containing Note This setting allows specification by a string Enter this information into the Content column Sample Type Sample Control Allelic Ladder Primer Focus or Negative Control Sample Type Sample oF Megqative Control 5 36 GeneMapper ID Software v3 1 User Guide Section 5 3 Table Settings Tab Table 5 10 Samples view filter settings continued Column Name Filter Settings Reference Data Off scale Sizing Quality Invalidated Yes or No Show All Records Show All Records Sample File Not Found Matrix Not Found Size Standard Not Found Show Pass green squares or Show Low Quality red octagons show All Rec Show All Records show Pass Show Low Quality Sizing Quality SQ Show Pass Show Check or Show Low Quality Show All Rec Mg Show All Records show Pass Show Check Show Low Quality Note These settings restrict the display to those records meeting these criteria Genotypes View Filter Settings Note In applying these settings to filter parameters be aware that when
296. r genotypes table shown in the Samples Plot window Look in GeneMapper x c FA fo File As Tab delimited text ixt hl Export Table Ba a Recent My Documents GM3 1 Database Defaults installer Size Standards Table Settings was B GeneMapper_Log a B PanellmportLog My Network File name Database Export Table Files of type frav and Comma delimited Text tt csv v Cancel Note This command is functional only when a table is currently being viewed The exported table contains the same configuration as the currently viewed table Close Plot Prompts to save any pending changes and closes the Window plot window Escape The Edit menu command provides basic editing for the Plot window menus described in this section Ondo CA A Edit menu item and description ltem Description Undo The Undo command is enabled after a user action that Ctrl Z adds modifies clears or reorganizes It allows you to undo certain actions such as e Editing an Allele e Editing a Bin e Resizing Markers e Zooming GeneMapper ID Software v3 1 User Guide Plot Window Menus View Menu The View menu contains features you can use to control e zooming of electropherograms e electropherogram lines dye displays on the view e anumber of display activities AXIS Scale p Tools Alleles Help Toolbar Sampl
297. r standard fragments that may run anomalously Use the Peak Detection parameter options to specify the minimum peak height to be detected for analysis This in turn controls the number of peaks analyzed Peaks falling below the parameters specified display in the electropherogram but are not analyzed and no values display for them in the tabular data Peak Detection Parameter Options Item Description For example Peak Amplitude Thresholds Set the dye amplitude threshold at a level that allows the software to detect peaks but eliminate noise For each dye the GeneMapper ID software detects peaks above the threshold entered in the entry field If you leave the default value of 50 peaks with amplitude above 50 are analyzed and display in the tabular data Lower amplitude peaks still display in the electropherogram but are not analyzed and do not display in the tabular data GeneMapper ID Software v3 1 User Guide B 9 Appendix B Software Genotyping Algorithms Split Peak Correction Parameter Options Item Description For example Minimum Peak Half Defines what If this number is large Width constitutes a peak the software ignores noise spikes Use to specify the smallest half peak If the peaks in the data width for peak are narrow set the detection value to a low number The range is 2 99 Experiment with this value to determine the
298. re 4 Export Table Exports the selected table button 5 File name field Enter a file name to use for the exported table 6 Show files of This is a display filter for files folders are type field always shown Use to select the file extensions for display e All files e txt files e csv files e txt amp csv 7 Cancel button Closes the Export Table window without exporting a table 8 Export file as Select the format of the file you want to field export Note Exported text tables show PQVs as numbers even if they are displayed as symbols GeneMapper ID Software v3 1 User Guide Project Window File Menu Dialog Boxes Print Dialog Box The Print dialog box shown below is a print file dialog box that controls printing The Samples table the Genotypes table Panel Manager views and plot windows may be printed i a pages Manm im Li r SEERon ltem Description Properties button Presents the dialog box used to set up the printer Name field Shows the currently selected printer Note The drop down menu to the right allows selection of other available printers Status Indicates status of selected printer Type Shows type of printer Where Shows the path name of the selected printer Comment Check box enables printing the project to a file Page range All is the only option available when working with tables Number of Select the number of copies to print
299. re shown Below is a representation of how the data for one sample will be printed when using this plot setting CO Control C mle vl x E E 100 200 200 400 CO_Conial COfle_yvl x CE 100 200 300 400 400 WE oo Renee iene sae ttm CO_Conirel Coma y1 x CE 100 200 300 400 1800 BOO To print this view 1 Within the samples tab of the main project window select the samples in the table you want to display in the printouts 2 Select Analysis gt Display Plots 10 8 GeneMapper ID Software v3 1 User Guide Printing Genotype Plots similar to Genotyper Using the Predefined AmpF amp TR Genotyping Setting 3 Select AmpFLSTR Genotyping from the Plot Setting drop down list Refer to Chapter 5 for information on importing plot settings into the GeneMapper Manager 4 Select File gt Print leaving all page set up options as default GeneMapper ID Software v3 1 User Guide 10 9 Chapter 10 Printing Data 10 10 GeneMapper ID Software v3 1 User Guide Process Quality Values This appendix describes how users can optimize the quality assurance level of their results by using the Process component based Quality Values PQV in the GeneMapper ID Software Version 3 1 to troubleshoot the data analysis process This appendix covers Overview of Process Component Based Quality Values PQV A 2 Using Process Com
300. re v3 1 User Guide l 19 Appendix GeneMapper ID Software Verification Matrix and Multi component Test I 20 pairs were tested to verify the non concordance feature in GeneMapper JD software using single source DNA samples inhibited DNA samples and degraded DNA samples All pairs were accurately defined with GeneMapper ID non concordance feature The fifth parameter evaluated in the Peak Detection and Genotyping category is the matrix and multi component test The same DS 32 and DS 33 matrix sample files were electrophoresed on the 310 Genetic Analyzers both Macintosh and Windows NT OS and analyzed with both GeneScan software v3 7 1 Windows NT OS and GeneScan software v 3 1 2 Macintosh OS Then these same sample files were analyzed by the Classic and Advanced algorithms in the GeneMapper JD software The mathematical matrices generated using the same scan data points show identical values Figures I 11 and I 12 lt GeneScan 3 7 File Edit Project Sample Settings View Windows Help H untitled Reactions B G i R ioo oa680 f oooz3 oooz4 foo025 Figure l 11 Matrix created in the GeneScan Software v3 7 GeneMapper ID Software v3 1 User Guide Results of Verification Testing Carte A Matrix Editor l x Matrix Description Matrix Mame STONT_Sdyes Description Matrix Settings Select the Matrix Standard Sample File Number of Dyes bal B Mo File Selected for B Data Start At 1000 gt Mo Fi
301. reate panels markers and bins for a SNaPshot Kit This feature can only be used with reference data generated from the ABI PRISM SNaPshot Primer Focus Kit Refer to the SNaPshot Tutorial for more information Vi ew M e n u Wj Eyi Full View Ctrl A AIS Scale Y Axis Scale Dyes Full View Depending on the X and Y axes mode the Full View command resets the X or Y axis to the e maximum range e panel size range e most recent Scale To range GeneMapper ID Software v3 1 User Guide 4 19 Chapter 4 Using Panel Manager 4 20 X Axis Scale for SNP genotyping data Scale to maximum Y Axis Scale t Scale to panel size range Dyes scale to The X Axis Scale menu command controls the horizontal scale of the reference data electropherogram being viewed in the Panel Manager as follows e Scale to maximum The electropherogram being viewed is scaled to its maximum base pair length as collected from the instrument e Scale to panel size range The electropherogram being viewed is scaled to the size range available for the selected panel e Scale to The user can define the horizontal range being viewed To zoom in to a specific region on the X axis of the electropherogram e Click and drag with the left mouse button the desired region on the X axis or e Right click on the X axis to open the X Axis Scale menu command options X Axis for microsatellite data male to maritim th
302. riginal location and the information contained within the GeneMapper project If you move the data from its original location this link is broken and the software will no longer be able to access the raw data to perform reanalysis If you want to reanalyze a project with a different copy of GeneMapper gain access to the raw and EPT data or if you have moved the data from its original location you must re establish the link Not only will re establishing the link allow you to reanalyze the project but it will also allow you to view the raw and EPT data Follow the steps below to re establish the link 1 Make sure the GeneMapper database you are importing the project into contains the same exact panels bins analysis method and size standard originally used to create the project If this is not the case export these items from the original copy of GeneMapper and import them into the GeneMapper copy you would like to view the data on Note This step must be done prior to importing the project 2 Export the project from the original GeneMapper database and import it into another GeneMapper database of choice These operations are done within the GeneMapper Manager window 3 Open the project from the File menu 4 View the Sample Info to determine the path between the project and the original sample file GeneMapper ID Software v3 1 User Guide F 3 Appendix F Transferring Data and Backing Up Data F GeneMapper ID 3 1 Un
303. rint Export Table Close Plotyvvindow Escape Cirl Shitt s Ctrl F Ctrl E File menu items and descriptions Command Description Save Panel Ctrl Shift S Opens the Save Panel s dialog box informing you of what panel s have been modified and how many projects currently in the GeneMapper D database will be affected The options are e Save Panel This option allows you to save any marker and bin changes made to the current panel name affecting all projects analyzed using this panel e Save Panel As This option allows you to save any marker and bin changes made to a new panel name affecting only the current project being analyzed Note When using Save Panel As feature the application creates a duplicate panel with bins from the current bin set and all bins from associated bin sets e Discard Changes This option allows you to discard any changes made to markers and bins Print Ctrl P Automatically prints the contents of the plot window in the following order e controls e samples genotypes e table if any Note Customize the view prior to printing GeneMapper ID Software v3 1 User Guide 8 5 Chapter 8 Using Plot Windows Samples and Genotypes 8 6 Edit Menu File menu items and descriptions continued Command Description Export Table Opens the Export Table dialog box allowing you to Ctrl E export either the sizing o
304. rogram by click drag on the X axis numbers Type the peak size in the corresponding column Note When you want to ignore a peak for the standard definition leave a zero in the Size column Press the tab key to move to the next size standard peak then enter its value Note Ensure you tab to the next size standard peak and not to a spike or to baseline noise See for all GeneScan size standard profiles and definitions Note To skip a fragment leave the value as zero in the corresponding cell in the table Click OK to save and close the Size Standard Editor or click Cancel to close without saving GeneMapper ID Software v3 1 User Guide Using the Size Standard Editor in Basic or Advanced Mode GeneMapper ID Software v3 1 User Guide Section 5 6 Size Standards Tab The Size Standard Editor window in Basic or Advanced mode allows you to define the peaks for each size standard dye you use Size Standard Editor Edit Size Standard Description Mama Description Size Standard Dye size Standard Table fd po ao eo spo po To ooo spo EEE OK Cancel To use the size standard editor in Basic or Advanced mode 1 Enter a name and description for the size standard you are creating 2 Choose a Size Standard Dye color from the drop down list 3 Enter the base pair values to be defined in your size standard in the table provided Note Use the Edit menu to undo
305. rv1 D138317 G f2 13 pppoe EU 24 Sample3 Databasing identifier_v1 D165539 G fs 11 in pin in nin ih mE 25 Sample4 Databasing Identifiler_v1 0165839 G 12 13 C E C C C E E a 26 Samples Databasing Identifiler_v1 D18551 Y 18 18 a C C C a E E E 27 Samples Databasing Identifiler_v1 0198433 Y 12 2 15 D C B C C E E E 283 Sample4 Databasing identifler_v1 D198433 Y f2 14 MEERE EEE EU 23 Sample5 Databasing identifierv1 D195433 Y f3 14 MEERE EEE EU 30 Sample3 Databasing Identifiler_v1 D21511 B 30 31 E C C C C E E a Ei Sample4 Databasing Identifiler_v1 D21511 B 28 29 C C C C C E E o 32 Samples Databasing Identifiler_v1 021811 B 29 30 DO C C C C E E a 33 Sample3 Databasing identifierv1 0381358 G fs 18 poppet E 34 Sample4 Databasing identifier_v1 381358 G f4 17 MEERE EEE EU 35 Sample5 Databasing Identifiler _v1 0381358 G 18 17 C C C C C E E ra 36 Sample3 Databasing Identifiler_v1 D5581 8 R 1 2 13 E E H 37 Sample5 Databasing Identifiler_v1 D55818 R 11 13 E E ra 3s Sample3 Databasing identifier _v1 D75620 B s 10 o EEG EU 393 Sample4 Databasing identifierv1 D75620 B j8 11 o EEG mU 40 Sample3 Databasing Identifiler_v1 pest 179 B 1 2 14 E E m 41 Sample4 Databasing Identifiler_v1 pest 179 B 1 0 16 E E a 42 Sample5 Databasing Identifiler_v1 D851179 B 12 13 E E a 43 Sample3 Databasing identifier_v1 FGA R fja 28 EEG EU 44 Sampled Databasing
306. s Information from the Data Collection Sample Sheet or Plate Record Introduction Transfer of Information Using Sample Sheet Information G 2 The GeneMapper ID software uses the sample sheet output from the ABI PRISM Data Collection instrument to identify the lane number and contents of each sample during analysis This section describes the sample sheet information used by the GeneMapper ID software so that you can correctly set up the sample sheet For detailed information on how to complete a sample sheet correctly see the user guide for the appropriate ABI PRISM instrument Data is transferred from a sample sheet into GeneMapper JD software as described in the following table Sample Sheet GeneMapper D Software Sample file name and Sample file name and Sample name Sample name Note These names are the same between a sample sheet and GeneMapper D software Sample Info or Color Info Sample type Control Allelic Ladder Sample Negative Control Color comment Comment if selected in the Samples View table Comment after Panel name Use User Defined Columns 1 3 the pipebar to separate text into user defined columns The sample sheet information is essential for associating the nature of sample fragments with individual dye lanes and tables in the GeneMapper JD software For example the following figures show how the GeneMapper ID software incorporates information entered in t
307. s Use the Slope Threshold for Peak Start and Slope Threshold for Peak End parameters to adjust the start and end points of a peak This parameter can be used to better position the start and end points of an asymmetrical peak or a poorly resolved shouldering peak to more accurately reflect the peak position and area In general from left to right the slope of a peak increases from the baseline up to the apex From the apex down to the baseline the slope becomes decreasingly negative until it returns to zero at the baseline Apex Increasingly Increasingly positive negative slope pew Baseline Q g If either of the slope values you have entered exceeds the slope of the peak being detected the software overrides your value and reverts to Zero As a guideline use a value of zero for typical or symmetrical peaks Select values other than zero to better reflect the beginning and end points of asymmetrical peaks A value of zero will not affect the sizing accuracy or precision for an asymmetrical peak GeneMapper ID Software v3 1 User Guide B 25 Appendix B Software Genotyping Algorithms Using These Use the table below to move the start or end point of a peak Parameters IF you want to move the THEN change the start point of a peak Slope Threshold for Peak Start value closer to its apex from zero to a positive number end point of a peak closer to its apex Slope Threshold for Peak
308. s 4 22 Panel data hierarchy 4 4 Panel definition format of text file 4 23 Panel definitions for SNaPshot 4 29 Panel Manager Add Bin command 4 13 Add Reference Data command 4 17 Auto Bin command 4 18 Auto Panel command 4 19 Bin view 4 40 4 41 Bins menu 4 12 Delete Bin command 4 14 Delete Bin Set command 4 17 Delete SNP Marker command 4 13 Duplicate Panel command 4 8 dyes 4 21 Edit Bin command 4 14 Edit Bin dialog box 4 15 Edit SNP Marker command 4 12 Full View command 4 19 Import Bins Options dialog box 4 9 Import Export Bin Set command 4 9 Import Export Panels command 4 8 Navigation pane 4 6 New Bin Set command 4 16 new marker 4 8 new panel 4 8 overview 4 2 Index 3 Panel Manager continued Panel SNP Reference Datacommand 4 18 purpose of 1 6 4 2 rules for imported and exported files 4 33 Show Project Alleles command 4 17 toolbar 4 5 View menu 4 19 window commands 4 7 X Axis Scale menu command 4 20 Y Axis Scale 4 21 Panel Manager window 4 3 Panel names 4 35 Panel table columns 4 35 Panel Table view 4 34 parameter values 5 16 Pass check or low quality PQV metrics 5 25 PCR Single stranded Conformation Polymorphism SSCP Minimum Peak Half width setting B 10 B 15 peak detection optimizing sensitivity B 20 B 22 B 24 parameters B 17 peak window size B 17 polynomial degree B 17 Slope threshold B 25 peak detection algorithm advanced 5 19 classic 5 21 Peak Detector tab advanced peak detection algori
309. s designed specifically for AmpF STR data analysis but it can perform other microsatellite and SNP analysis and this user guide includes information needed for microsatellite and SNP analysis To obtain more information about the initial setup and workflow specific to a particular application refer to one of the three tutorials listed in Related Documentation on page x The GeneMapper ID Software Version 3 1 has not undergone specific developmental validation for human identification HID applications Human identification laboratories that choose to use GeneMapper JD Software for data analysis should perform their own developmental verification studies The GeneMapper JD software v3 1 supports data generated from the following instruments e ABI PRISM 310 Genetic Analyzer e ABI PRISM 377 DNA Sequencer e ABI PRISM 3100 Avant Genetic Analyzer e ABI PRISM 3100 Genetic Analyzer Note Data generated from a 310 Genetic Analyzer or 377 DNA Sequencer using a Macintosh platform must be converted to a Microsoft Windows based format before it can be used with the GeneMapper ID software Refer to Appendix E Converting Macintosh Sample Files GeneMapper ID Software v3 1 User Guide Overview of GeneMapper ID Software Supported The GeneMapper JD software v3 1 is specifically designed to work Chemistry Kits with the following AmpF STR kits and reagents AmpF STR Identifiler PCR Amplification Kit AmpF
310. s genotype concordance between overlapping loci among different AmpFZSTR kits for the same sample s or concordance of genotype calls from duplicate amplifications or duplicate injections of the same sample e Positive and negative controls give the expected allele calls 1 4 GeneMapper ID Software v3 1 User Guide Steps Required to Use GeneMapper ID Software Steps Required to Use GeneMapper D Software Using The following steps are required to use the GeneMapper JD software GeneMapper D You will be guided through this process in more detail by reading the Software GeneMapper ID Software v3 1 Human Identification Analysis Tutorial P N 4335523 Microsatellite Analysis with ABI PRISM GeneMapper Software v3 0 Tutorial P N 4335525 and SNP Genotyping with ABI PRISM GeneMapper Software v3 0 Tutorial P N 4335524 Set up GeneMapper ID Software Set up Project Window for Analysis e Set up the GeneMapper JD software as described below Import panels and bins in the Panel Manager Create an analysis method suitable for your application with appropriate bin sets Define custom views of analysis tables and plots View and set HID analysis options Create the appropriate size standard s 1f your data uses size standards other than those provided with GeneMapper D software Convert any Macintosh generated GeneScan sample files to the fsa format See Appendix E Converting Macintosh Sa
311. s were quantified using the QuantiBlot Human DNA Quantitation kit and amplified with the AmpF STR kits Note For each sample type all AmpF STR kits used the sample quantity of DNA except the Identifiler kit The sample quantity of DNA for the Identifiler kit is denoted in parenthesis e Single Source DNA Samples Eight female and eight male DNA samples were amplified with 2 ng total input DNA 1 ng for the Identifiler kit e Mixture Samples Three mixtures of male and female DNA were amplified per kit These samples were mixed in defined ratios of 1 to 9 1 to 1 and 9 to 1 respectively e High Quantity DNA Samples Three DNA samples were amplified with 8 ng total input DNA 4 ng for the Identifiler kit e Low Quantity DNA Samples Three DNA samples were amplified with 250 pg total input DNA 125 pg for the Identifiler kit e Degraded DNA Samples Three DNA samples were degraded with 2 units DNase I for 15 minutes with subsequent amplifications using 4 ng uL degraded DNA 2 ng uL for the Identifiler kit e Inhibited DNA Samples Three 2 ng uL total input DNA samples were amplified in the presence of 16 uM hematin 1 ng L for the Identifiler kit The samples were amplified using an Applied Biosystems GeneAmp 9700 PCR system and the recommended cycling conditions The amplified samples were injected or loaded on three or more ABI PRISM instrument platforms GeneMapper ID Software v3 1 User Guide
312. samples results in failed analysis Allelic bin definitions are stored in the AmpF STR panels in the Panel Manager Lanes or injections containing the allelic ladder should be analyzed with the same analysis method and parameters used for samples GeneMapper ID Software v3 1 User Guide Analyzing Samples Files e Alleles not found in the AmpF STR Allelic Ladders do exist These off ladder alleles may contain full and or partial repeat units An off ladder allele is defined as an allele falling outside of the 0 5 bp bin window of any known allelic ladder allele or virtual bin Note Ifa sample allele peak is called as an off ladder allele then the sample needs to be run again to verify the result e The marker specific stutter ratios included in the kit panels and cutoff value used in the tutorial are provided to serve as a tool and a guideline Final conclusions should be based on careful examination of the STR profiles Analyzing Samples Files Sample File Format To analyze sample files you can either add fsa files generated on an Applied Biosystems Genetic Analyzer or you can open an existing project apply new analysis settings and re analyze IMPORTANT In order to be able to re analyze an existing project certain criteria must be met Please see Appendix F for specific information regarding re analyzing existing projects GeneMapper JD Software v3 1 supports data sample files generated from the following ins
313. scending Z A 3 2 1 4 Click OK The table will sort automatically after clicking OK 7 28 GeneMapper ID Software v3 1 User Guide Genotypes Table Find Option Ifyou are looking for a particular sample or a particular set of samples you can use the find option To use the Find option 1 Ifthe genotypes table is not already displayed click the genotypes tab in the main project window or select View gt Genotypes 2 Select Edit gt Find Find what In Column Sample File r Fing Mext Find ll Close 3 Enter the string of text or numbers you wish to find and select what column you want the software to look in 4 Click Find Next to find the first instance or click Find All to find all instances at once The rows containing this item are highlighted in the genotypes tab Genotypes Plots When you have a row s highlighted you can display genotypes plots Genotypes plots display the peaks for a particular marker loci Highlight the desired rows and select Analysis gt Display Plots Refer to Chapter 8 for more information about viewing genotype plots GeneMapper ID Software v3 1 User Guide 7 29 Chapter 7 Analyzing and Interpreting Data Interpreting Genotype PQVs and Genotype Quality Genotype PQVs 7 30 Genotype PQVs and Genotype Quality are displayed in the Genotypes Table and Plots To view the Genotype Table click the Genotypes Tab from the main project window Or if you have
314. scription Automatic This option button sets the level automatically for option button the five dye colors blue green yellow red and orange This level represents the minimum signal strength that will be identified as a peak for each dye equivalent to 10 times the noise User specified rfu This option button enables the number entry fields option button for the five dye colors blue green yellow red and orange These numbers are the minimum signal strength that will be identified as a peak in relative fluorescent units rfu For all dyes the default is 50 the minimum is 1 and the maximum is any number of 10 digits B 6 GeneMapper ID Software v3 1 User Guide Classic Mode Peak Detection Algorithm Settings Classic Mode Peak Detection Algorithm Settings Overview In Classic mode there are five analysis parameter options They are as follows Ranges Data Processing Size Calling Method Peak Detection Split Peak Correction Ranges The following are the Ranges parameter options Parameter Options Item Description Analysis Used to analyze all the data collected on the drop down menu genetic analysis instrument for each sample Full Range Enter Start and Stop data point numbers in the Partial Range entry fields in order to specify only a limited range analyzed for each sample The data point numbers affect what is displayed in the results display Normally set the analys
315. se errors alerts occur How many installation attempts have you tried Are you logged in to the local computer Is the TCP IP configured Do you have administrative privileges Is this a stand alone computer networked or attached to an ABIP s Instrument H 2 GeneMapper ID Software v3 1 User Guide Troubleshooting Sheets For Software Related Issues Does the computer and operating system meet the required computer specifications List any potential discrepancies Problem description Did you receive any error messages or alerts If so what did they say if possible take screen captures At what point did these errors alerts occur Is the problem reproducible Steps taken prior to encountering or to reproduce the problem i GeneMapper ID Software v3 1 User Guide H 3 Appendix H Troubleshooting Sheet For Low Quality Collect the following files and complete the questions below Analysis 7 2 3 Sample files fsa files imported into GeneMapper ID Analysis Method export of GeneMapper Manager Size Standard Definition if different than default export out of GeneMapper Manager Matrix File 310 377 Only export out of GeneMapper Manager Panels and Binset if different than default export out of Panel Manager Does the computer and operating system meet the required computer specifications List any potential discrepancies Problem description
316. selected peak The label is then deleted If the Show type of edit checkbox is selected in the plot profile the word Removed displays inside the label box if Allele Changes is selected from the View menu Note Multiple alleles may be deleted at the same time by selecting multiple peaks This is constrained to the peaks in the plots currently shown History Allows you to view the allele history in columns labeled Basepair Allele Name User Name Modification Date Action and Comments GeneMapper ID Software v3 1 User Guide 8 25 Chapter 8 Using Plot Windows Samples and Genotypes 8 26 Alleles menu items and descriptions continued Item Description Rename Allele Rename Allele Allele Edit Comment Ctrl 0 Add Bin Ctrl B Edit Bin Ctri U Delete Bin Delete g 10 11 12 13 14 15 7 D 4a amp M Custam Allows you to change the allele call on a labeled peak 1 Select Peak Selection mode 2 Select a labeled peak using the left mouse button 3 Select the Rename Allele menu item or click with the right mouse button to open the allele edit options and click Rename Allele 4 Select an allele call from the menu to rename the allele If you select Allele Edit Comment the Edit Allele Comment dialog box opens allowing you to enter an allele comment for the selected peak The allele is then renamed to the selected allele call lf Show type of edit prefix is t
317. standard peak or click OK if you are finished applying your changes to the edited peak GeneMapper ID Software v3 1 User Guide Size Calling Curve Size Calling Curve Overview The Size Calling Curve is used as a measure of how well the internal size standard matches the standard definition and whether or not it is linear The Size Calling Curve displays two curves F Size Match Editor X File Edit View Tools XB S Size Matches Size Calling Curve PP_Sample2 tsa Best Fit Second Order Curre AD 242 ORS4R S05 TORTS AI 0 080095376062 12441 AG 4 15105 39504396385 6 6 R z 0 9999942020153115 Size Calling Curre Local Southem wiethod 4000 S000 6000 DE Cancel Apply Red Curve Ifthe size calling curve is a red curve it is based on the size calling method used to analyze the data Black Curve Ifthe size calling curve is a black curve it is a best fit least squares curve which the GeneMapper JD software calculates for all samples regardless of the size calling method used This curve is provided to help evaluate the linearity of the sizing curve When the sizing curve and best fit curve match they overlap so you see only the size curve Note Sizing problems due to anomalous mobilities may be displayed as non linear Sizing Precision Since the 250 bp peak is not defined when running on capillary using the 250 bp electrophoresis platforms an analyst can use this peak to check the peak of GS 50
318. ta Point Ctri N GeneMapper ID Software v3 1 User Guide 8 9 Chapter 8 Using Plot Windows Samples and Genotypes View menu items and descriptions continued ltem Description Y Axis Controls the vertical scale of the electropherograms Scale currently viewed as follows e Scale Individually Each electropherogram being viewed is scaled to its maximum height e Scale To Maximum Y All plots currently viewed are scaled to a global maximum value e Scale To This setting allows a user to set the Y Axis scale value If multiple plots are currently viewed all plots are scaled to this value Scale Individually Labels Scale To Maximurn r Bins Ciri B Scale To Zooms in on a specific region on the Y axis of the electropherogram as follows e Click and drag with the left mouse button the desired region on the Y axis This action only affects the selected electropherogram or e Right click the Y axis to open the Zoom To dialog box which enables specific zoom coordinates and tick spacing units to be set Use the Apply To checkbox to apply the settings to all electropherograms panes F Y Axis Zooming TAXIS zoom To Tick Spacing unitsitick Apply to all electropherogram Cancel e Double click on the Y Axis to return selected electropherograms to Full View e Shift double click on the Y Axis to return all electropherograms to Full View
319. tabasing Identifiler_v1 THO1 G 9 9 3 E B amp ca B a a B ia 71 Sample4 Databasing Identifiler _v1 D135317 G 12 13 E E E EEEE E E 72 Sample4 Databasing Identifiler_v1 D165539 G 12 13 E E EE EEEE E E 73 Sample4 Databasing Identifiler_ 1 D251338 G 25 A BE E fa E E g 74 Sample4 Databasing Identifiler_v1 D195433 Y 12 14 E E E a E E eB a 75 Sample4 Databasing Identifiler_ 1 AJA 16 18 E ra E E a ica i ga E 76 Sample4 Databasing Identifiler_v1 TPOX Y 8 H E E m ic a E E 77 Sample4 Databasing Identifiler_ 1 D18551 Y f8 a E E B E E B E 78 Sample4 Databasing Identifiler_ 1 AMEL R x E o E G w A B H 79 Sample4 Databasing Identifiler_ 1 D55818 R 12 ia m E m E E Ei Ei 60 Sample4 Databasing Identifiler_ v1 FGA R 21 26 E Bg Bi ge E E E m H 51 Samples Databasing Identifiler_ 1 DBS1179 B 12 13 E E E E TI B E B E 82 Samples Databasing Identifiler_v1 D21511 B 129 30 E E E E B E E E 83 Samples Databasing Identifiler_v1 D75820 B 11 Es E a cal a w iP a Samples Databasing Identifiler_ 1 B 10 A M B E 7 M M 5 85 Samples Databasing Identifiler_ 1 D3S1358 G 115 17 6 E E ja iar a ita E iat 86 Samples Databasing Identifiler_v1 THO1 G 16 9 3 a E E E E E EA a a oF Samples Databasing Identifiler_v1 D135317 G 11 C a a al E E E i 58 Samples Databasing Identifiler_v1 D16S539 G 111 E rai E E m a BE iH 6g Samples Databasing Identifiler_v1 D2513368 G 23 a ial E Bi a ied ie E 90 Samples Databasing Identifiler_v1 D195433 Y 13 14 w E a a A E eal iz E 91 Sam
320. taskbar or to the title bar e Maximize the window to fill the entire screen e Resize the window by dragging any window edge or corner Reformatting the You can reformat the Project window Window e Use the Show Navigator switch on the View menu to show hide the navigation pane for the Samples and Genotypes views When the navigation pane is hidden the table occupies the entire width of the Project window e Shift click a column header to sort the data by that column The Sort dialog box under the Edit menu allows cumulative sorting over three columns see Sort Dialog Box on page C 46 e The first column of the table displaying row numbers is fixed during horizontal scrolling of the Samples and Genotypes tables e Create Hide Show columns as needed with different table settings created using the Table Settings in the GeneMapper Manager window e Resize columns by dragging the separating lines During resizing the cursor changes to a resize cursor a double headed arrow that looks like this lt 3 GeneMapper ID Software v3 1 User Guide 3 3 Chapter 3 Using the Project Window 1 2 A26 SAMM 3 Project Samples Genotypes _ Databasin Status Sample F Sample MN Sample IC Comment Sample T Specimer Analysis Panel Size Star Matrix Run Nar Instrumer lInstrumer Run Date ID_Contre ID_Contre None Pampie Jro expon HID_Adve Identifiler CE_G5_F Databasit ABI3100 demo_31 2002 06 4 ID_Ladde ID_Ladde None Allelic Laino
321. ted on the whole height of the page on a printer C 36 GeneMapper ID Software v3 1 User Guide Add Samples to Project Dialog Add Samples to Project Dialog Introduction This section provides general information about e the Add Samples to Project dialog e the Edit and View menus available in the window e using the three tabs in the window This following topics are covered in this section Topic See Page General Information and Procedure C 37 Edit Menu C 38 View Menu C 40 Files Tab C 41 GM Database Tab C 42 General The Add Samples to Project window provides a navigation pane to Information and enable you to add Samples to the project You can also access the Procedure dialog box by clicking the Add Sample icon on the Project window toolbar Add Samples to Project _ 4 xl Edit View Files Gh Database Samples To Add l n atabasing a Microsatellite E SMaPshot CJ GM34 Database Defaults 0 JRE 9 Panels L Plot Settings 9 Size Standards C Table Settings By app Add To List gt Clear z Add TEA Cancell Options GeneMapper ID Software v3 1 User Guide C 37 Appendix C Project Window Software Interface To add samples to the Project 1 Click Add Samples To Project on the Project window toolbar Add Samples to Project window opens to the last tab viewed by you default is Files tab Sample files can reside on the local hard drive s mapped network
322. the samples 1 e the PQV Results The ability to display plots for the 20 364 alleles evaluated in these various views 1s 100 accessible and functional The second parameter tested in the Workflow category 1s the ability to display default and customized plot settings A user can create and edit plot settings For example a trained analyst wanting to interpret a mixture sample can create a plot setting to view electropherograms in only one color Six projects were evaluated for the functionality of the plot settings to display the data as defined in the plot settings editor Five different predefined display settings were evaluated AmpF STR sizing AmpF STR genotyping Overlay ROX Overlay LIZ and customized plot settings displays only one color at a time Results The ability to display data as defined in the plot settings editor is 100 functional with the six projects tested The third parameter tested in the Workflow category is the printing test GeneMapper ID Software v3 1 User Guide Results of Verification Testing In the GeneMapper D software various plot performance tests were conducted These tests measure performance attributes such as time to display plots time to scroll through plots and sample configurations The result for the amount of time to display sample configurations e g display different panes samples and dyes was less than ten seconds In the GeneMapper JD software the results for vi
323. the cell is closed the software must check that the name is unique An alert message will display if the name is not unique Comments Contains comments regarding the panel Cell accepts alphanumeric characters Panel Name Rule Panel names must be unique within a kit and across kits GeneMapper ID Software v3 1 User Guide 4 35 Chapter 4 Using Panel Manager Marker Table View Introduction Selecting a panel in the navigation pane displays the Marker table for that panel The Marker table shown below enables you to view create and edit marker names and associated data EA F Panel Manager File Edit Bins View HE E e e E E Panel Manager Maker Hame Dye Color Win Sie Max Sie Cortrol Alleles Marker F Marker Speci Comments Ladder Alleles E Bin set S ao ToC el S cotiter_v1 5 D3si3s green a80 faso fisas o fja fon hone 243441541617 6 7 5 g El AmpFLSTR_Panels_v1 H EE w E E Green w H E SGM Plus v1 THOM green 1 59 0 205 0 893 4 0 05 none 4 5 6 7 9 9 3 10 ia Easetier vi D1 35317 green 205 85 2501 6 I 1 4 0 08 none jaa 041142434 H f Protiler Plus _CODIS D1 65539 green 2553 301 A I 1412 4 a1 none 58 31 041 12 13 H E COtier eons vi D251 338 green 304 6 370 31 1 923 4 a1 1 none 15 16 17 1819 2 identifier coDis_v1 4g D1 95433 yellow 101 D 148 0 14 15 4 0 13 none 910 11 1212 21 7 l 11 ANA yelow 151 D BEE 4716 4 013 none 11 12 13 14 15
324. the size calling curve To open the Size Match Editor window click Analysis on the toolbar in the GeneMapper JD project window and select Size Match Editor Psi i E Size Match Editor ae X File Edit View Tools x Eg PP_Sarmple1 tsa size Matches Size Calling Curve PP_Sample2 tsa Sizing Quality lt 1 0 gt Gweride SG OK Cancel Apply When the Sizing Quality is a low value you can use the Size Match Editor to adjust the in lane size standard Typical problems include the following conditions e Peak shift The algorithm does not correctly identify a peak resulting in invalid positions for other size standard peaks e Missing peak The algorithm does not identify a peak e Extra peak The algorithm identifies a peak that is not a size standard peak but a spike or pull up peak 7 14 GeneMapper ID Software v3 1 User Guide Size Match Editor Window The Size Match Editor window elements are described in the Elements following table below Element Description Sample navigation pane Enables you to select a sample to view by scrolling through the Sample list The list of samples displayed reflects the list of samples selected in the Project Sample view Size Quality indicator Displays the quality of the currently displayed size standard name listed above the plot Value 0 0 1 0 Size Matches electropherogram Displays the size standard peaks Clicking a peak selects it s
325. thm 5 19 basic peak detection algorithm 5 19 classic peak detection algorithm 5 21 Peak Quality tab allele number 5 23 heterozygote balance 5 23 peak morphology 5 23 pull up peak 5 23 signal level 5 23 SNP 5 23 plot settings default profiles 5 41 Plot Settings Editor Display Settings tab 5 50 General tab 5 43 Genotype Header tab 5 45 Labels tab 5 47 Index 4 Sample Header tab 5 44 Sizing Table tab 5 46 plot settings editor 5 43 Plot window Add Allele option 8 25 Add Bin option 8 27 Allele Changes option 8 15 Allele Edit Comment option 8 26 Alleles menu 8 23 bins 8 13 Controls To Top option 8 15 customizing 8 3 Delete Allele option 8 25 Delete Bin option 8 28 displaying 8 2 Dyes option 8 16 Edit Bin option 8 27 Edit menu 8 6 Editing Mode option 8 23 example of 3 14 Export Table dialog box 8 6 File menu 8 5 Full View option 8 8 Header option 8 20 Help menu 8 28 History option 8 25 labels 8 11 Marker Indicator option 8 21 Marker Margin option 8 21 Marker Range option 8 20 menus 8 5 Off scale Peak Indicator option 8 16 overview 8 2 Peak Positions indicators 8 13 Plot Settings option 8 22 Plots option 8 17 Rename Allele option 8 26 Table Filter option 8 19 Tables option 8 17 8 19 toolbars 8 4 Tools menu 8 22 View menu 8 7 X Axis Scale option 8 9 Y Axis Scale option 8 10 PlusA Distance 5 18 PlusA Ratio 5 18 GeneMapper ID Software v3 1 User Guide polynomial degree peak detection B 17 varying B 18 w
326. ting depends on the memory specifications of your printer e In GeneMapper ID software various print performance tests were conducted These tests measured the total time mm ss needed to print a one page document under various plot configurations Print time was measured by time taken to spool document plus time for spool to leave printer queue Time did not include the time taken for the printed page to exit the printer Differences in time can result if the document is printed on a slower CPU 1000 MHz versus a faster CPU 2386 MHz and if the GeneMapper ID CPU had less RAM The printer used for comparisons in house was an HP 4500 e Applied Biosystems cannot test all available printers on the market and therefore cannot guarantee printing will be successful with each and every printer available GeneMapper ID Software v3 1 User Guide 10 3 Chapter 10 Printing Data Page Set Up Options Table Tab 10 4 Options Page Setup x Font Times Nev Roman 7 S ZE fio m Use Screen Font Page Order Print column first Print rows first i Print All Data Start Mew Page Page Setup OK Cancel e Page Order When tables are printed the number of columns and rows can exceed the limit so that they cannot be printed on one page Example A table has 50 columns and 10 rows A page can accommodate 10 columns only so five pages are required to print 50 columns horizontally Similarly if t
327. ting in the detection of a third peak Peak C In these two observations the filtering in Genotyper software compared Peak A to Peak C The GeneMapper ID software appropriately filters the stutter and the shoulder peak using defined bin sets EA Genotyper 3 7 Plots 3100NT_PROFILER_PLUS_original gta File Edit Analysis Category Table Views Macro Window Help 180 182 184 186 155 190 192 194 196 198 200 202 2 _ AO Nele_4 3ng 02 fsa 2 Green fele_4 3ng 311 108 igri Figure l 8 Genotyper software v3 7 GeneMapper ID Software v3 1 User Guide l 17 Appendix GeneMapper ID Software Verification Editing Functionality Test I 18 E Samples Plot File Edit view Tools Alleles Help Plot Setting HID Genotyping p Panes 3 EE ala E Pa MB Le fi T ah A Sample Name Pand s90 sQ Figure l 9 GeneMapper ID Software v3 1 using the Advanced algorithm The third parameter evaluated in the Peak Detection and Genotyping category is the editing functionality test This parameter was evaluated with 500 alleles for the ability to change allele calls view the history of the allele calls and display the changed allele calls These abilities functioned with a 100 success rate If multiple alleles have been edited as shown in Figures I 10 the history can be displayed for each edited peak by selecting the peak and displaying the history If an allele is edited more than once then only the last edit is
328. tion software v2 0 GeneMapper ID Software v3 1 User Guide 5 65 Chapter 5 Using GeneMapper Manager Size Standards Table 5 16 Description of elements in the Size Standards tab Tab Elements 5 66 Item Description Name column Size standard name Last Saved column Owner column Date Time stamp showing when the size standard was last saved User name of the person who created the size standard Type Classic or Basic Advanced Description A description of the size standard New button Opens the Specify parameters and Size Standard Editor dialog boxes to create a new size standard Open button Opens the editor for a selected size standard Enabled when a single size standard is selected Save As button Displays the Save As dialog box Enabled when a single size standard is selected Import button Displays a dialog box for Importing size standards Always enabled Export button Displays a dialog box for Exporting selected size standard s Enabled when one or more size standards are selected Delete button Deletes the selected size standard s Done button Closes the GeneMapper Manager GeneMapper ID Software v3 1 User Guide Section 5 6 Size Standards Tab Creating Editing a Size Standard Overview Specifying Size Standard Parameters To create a size standard you must first specify the parameters under which the size standard is created
329. tions as shown in the following table Case Result A sample file field contains The sample type for the sample is set either Control Ctrl Cri Cntrl to Control or a similar variation Analysis Tab The Analysis tab contains preferences associated with data analysis Startup Add Samples Analysis ma Automatic Analysis i Automatically bring low quality samples to the top Guality Metrics Display Symbols C Numbers If only one labelled allele ina genotype then duplicate the label Duplicate homozygous alleles OK Cancel GeneMapper ID Software v3 1 User Guide 6 7 Chapter 6 Options Dialog Box 6 8 Element Description Automatic Analysis group box Automatically brings errors to the top of the table check box When samples are analyzed the software sorts the table automatically to bring samples with errors to the top of the table Same functionality as the Bring Errors To Top icon in the Project window Quality Metrics Display group box Symbols button Numbers button Display quality values in project tables as symbols green square yellow triangle red octagon or numerical values 0 0 1 0 Note If Symbols default is chosen then all quality values are shown as symbols If Numbers is chosen then only SQ and GQ columns are shown as numbers the other result columns are symbols If only one allele is labeled in a genotype
330. tions on how to use the software refer to the GeneMapper ID Software v3 1 Human Identification Analysis Tutorial PN 4335523 Audience _ This guide is intended for GeneMapper JD software users Assumptions This guide assumes that you have read all applicable material and installed the GeneMapper ID Software Version 3 1 according to the installation instructions included in the software kit This guide uses conventions and terminology that assume a working knowledge of the Windows operating system the Internet and Internet based browsers Text Conventions This guide uses the following conventions Bold indicates user action For example Type 0 then press Enter for each of the remaining fields e Italic text indicates new or important words and is also used for emphasis For example Before analyzing always select the appropriate size standard e A right arrow bracket gt separates successive commands you select from a drop down or shortcut menu For example Select File gt Open gt Project GeneMapper ID Software v3 1 User Guide ix Preface User Attention Words Two user attention words appear in Applied Biosystems user documentation Each word implies a particular level of observation or action as described below Note Provides information that may be of interest or help but is not critical to the use of the product IMPORTANT Provides information that is necessary for proper use of the sof
331. titled gmid Is Logged In Fie Edit Analysis View Tools Help e co Sy OE tate setina o Tawe Eli Project nfo Raw Data EPT Data Casework gO Control tse Sample Information arre CO Ladder fsa sample File CO Sample tsa sample Origin Path saa CO Sample tsa E ppliedbiocsystenms GeneMapper Data CaseworkCO Samplel tsa a PP Cortrol fsa tatus Message Need to be Analyeec A see PP Ladder fsa File Source Disk media H BP Sarnpled fsa bi Progress Status Os i Pathway to the samples used in this project 5 Create or re create this path You must use the exact same file names in order to successfully establish the link In the example above you need to create the exact same folders Data and Casework within E AppliedBiosystems GeneMapper a In the GeneMapper folder create a new folder titled Data GeneMapper File Edit View Favorites Tools Help da Back ob G Search Dh Folders History r Select an item to wiew its SeratSebase description aa docs See also _ 6M3 1 Database Defaults My Documents l installer My Network Places I JRE C Panel My Computer ANS z 24 objectis 593 KB My Computer a GeneMapper ID Software v3 1 User Guide Transferring Data b In the Data folder create a new folder titled Casework File Edit View Favorites Tools Help da Back G Search C Folders History r Casework Select
332. to add the panels and bins back into your analysis methods Therefore it is best to make a copy of the panel before you modify it Note Applied Biosystems has optimized the panels and bins provided for AmpF STR data analysis Therefore Applied Biosystems does not recommend changing the marker information If this information is changed Applied Biosystems requests that you analyze with the default values if applications support is needed GeneMapper ID Software v3 1 User Guide Bins View Microsatellites Only Bins View Microsatellites Only Note The information contained in this section Bins View is not relevant to AmpF STR data analysis Introduction The Bins view enables you to view create and edit bins The data F Panel Manager displayed in the Bins view imported into the Panel Manager using the Import Bin Set command is used to create the bins associated with the marker Bins allows you to average the minor variations in size that occur from run to run and capillary to capillary File Edit Bins View E X E E Bin Set ampFLSTR_Bins_v1 Tea igg ole gt Profiler _Pi A D351358 D21511 D18551 D55818 H D135317 w 4 b Reference Samples 08 0 6 0 5 0 4 0 3 0 2 0 1 14417 1214 125 129 133 137 1 145 149 153 157 161 165 D851179 4 b X 164 02 Y 0 49 Bin 17 163 50 164 50 Marker D851179 118 00 183 50 OK Cancel Apply GeneMappe
333. to update the navigator pane Duplicate Panel The Duplicate Panel command creates a copy of a selected panel and places it in the same kit as the original with the name Selected Panel dup For example the duplicate name for the New Panel is New Panel dup The panel and all associated bins with all bin sets are duplicated as well This menu item is enabled when a panel folder is selected in the navigation pane Import Export Panels Refer to the GeneMapper ID Software v3 1 Human Identification Analysis Tutorial PN 4335523 for more information on importing panels and bin sets specific to the AmpF STR PCR Amplification kits The Import Panels command and the Export Panels command open the corresponding dialog box to allow importing exporting of kits and panels This command is enabled when you select the Panel Manager root node in the navigation pane See Formats of Panel and Bin Text Files on page 4 22 for more information GeneMapper ID Software v3 1 User Guide Panel Manager Window Commands Export Panels is not available when the Panel Manager root node 1s selected it is only active when a kit is selected Import Export Bin Set The Import Bin Set command and the Export Bin Set command open the corresponding dialog box to allow importing exporting of bin sets This command is enabled when you select a kit name in the navigation pane See Formats of Panel and Bin Text Files on page 4 22 for more in
334. truments e ABI PRISM 310 Genetic Analyzer e ABI PRISM 377 DNA Sequencer e ABI PRISM 3100 Avant Genetic Analyzer e ABI PRISM 3100 Genetic Analyzer The fsa file extension is required for GeneMapper ID software to recognize the file Sample files created from an instruments using a computer running Microsoft Windows NT or 2000 operating system automatically create file with this extension Note Data generated from a 310 Genetic Analyzer or 377 DNA Sequencer using a Macintosh platform must be converted to a Microsoft Windows based format before it can be used with GeneMapper ID software Refer to Appendix E Converting Macintosh Sample Files GeneMapper ID Software v3 1 User Guide 7 3 Chapter 7 Analyzing and Interpreting Data Adding Sample To add sample files to a new project Files to a New 1 Select File gt New Project to open a new project Project 2 Select File gt Add Samples to Project F GeneMapper ID 3 1 Untitled gmid Is Logged In Fie Edit Analysis View Tools Help New Project Ctrl h Open Project Chrl 0 Save Project Chrl 5 A sll ee Table Setting Save Project 45 4dd Samples to Project Ctril k Export Table Cbri E Export Table For GODIS Page Setup es es Erim Ctrl P ns es Log Out Alt F4 Exit 3 In the Add Samples to Project dialog box a Locate your sample files on your hard
335. tware instrument operation accurate chemistry kit use or safe use of a chemical How to Obtain More Information Related Documentation This GeneMapper ID Software Version 3 1 User Guide describes the GeneMapper ID Software Version 3 1 and provides information about the GeneMapper JD software for general use and customization The software comes with additional documentation to assist with installation and provide recommended workflows for specific genotyping applications The following are related documents e GeneMapper ID Software v3 1 Human Identification Analysis Tutorial PN 4335523 e Microsatellite Analysis with ABI PRISM GeneMapper Software v3 0 Tutorial PN 4335525 SNP Genotyping with ABI PRISM GeneMapper Software v3 0 Tutorial PN 4335524 e Installation ReadMe txt file located on the software CD IMPORTANT You must read the installation instructions and the ReadMe file before installing the software GeneMapper ID Software v3 1 User Guide How to Obtain Support Send Us Your Applied Biosystems welcomes your comments and suggestions for Comments improving its user documents You can e mail your comments to techpubs appliedbiosystems com How to Obtain Support Applied For the latest support information for all locations go to Biosystems http www appliedbiosystems com then click the link for Web Site Support At the Support page you can Search through frequently asked qu
336. ty value PQV An analyst can check the size quality of each sample by highlighting the individual samples in the navigation pane Further after evaluating each size standard an analyst may override the size quality value to 1 0 to confirm that each size standard was examined F Size Match Editor X File Edit View Tools xX Egl Al Female_28 13 Size Matches size calling Curve AZ Male_4 fsa A3 Male 23 2ng fsa sizing Quality 0 5 44 offscale_18 fsa A5 degraded _24 fsa AB inhibited 45 fsa Bi Female_30_2ng fza B2 Male_6 2ng fsa Ba 37 _22 1to9 fsa B4 offscale_33_8ng fsa BS degraded _24 fsa B6 inhibited _45 fsa C1 Female_31_2ng fsa Ca 37 _22 1tod fsa C4 offscale_38_8ng fsa CS degraded_kib2 fsa D1 Female_32_2ng fsa D2 Male_8_2ng fsa oon D2 7s En dta fom PoE Ok Cancel Apply Figure l 13 Size Match Editor using the Classic algorithm displaying a sizing quality value of 0 5 l 22 GeneMapper ID Software v3 1 User Guide Results of Verification Testing F Size Match Editor E x File Edit View Tools x Egl Ad Female_26 fsa Size Matches size calling Curve AZ Male_4 fsa A3 Male_23_2nq tsa Sizing Quality 0 9 44 offscale_18 fsa A5 degraded 24 fsa AB inhibited 45 fsa Bi Female _30_2ng fsa B2 Male_6 2ng tsa Ba 37 _ 22 1to9 fsa B4 offscale_33_8ng fsa BS degraded 24 fsa B6 inhibited 45 fsa C1 Female_31_2ng fsa Ca 3 _ 22 1tod fsa C4 offscale_38_8ng tsa C5
337. u command options a Blue Dye cCtri 1 Y Show SMP Bins Y Green Dye Ctrl 2 Yelow Dye Ctri 3 Red Dye Ctri 4 Orange Dye Ctri 5 w All Dyes Cirl 6 The Dyes menu item controls the dye colors shown when viewing SNP reference data and microsatellite reference data electropherograms in the Panel Manager Plot view checked when active You may select multiple dye colors GeneMapper ID Software v3 1 User Guide 4 21 Chapter 4 Using Panel Manager Formats of Panel and Bin Text Files This section provides an example of a file used to import panel definitions and a file used to import bin definitions These files are created using Microsoft Excel or other spreadsheet programs Files must be saved as a tab delimited text file Topic See page Panel Definition Example 4 23 Bins Definition Example 4 27 Panel Definitions for SNP Genotyping 4 29 Bins Definitions for SNP Genotyping 4 30 Import and Export Files 4 33 4 22 GeneMapper ID Software v3 1 User Guide Formats of Panel and Bin Text Files Panel Definition The following examples show panels defined in the format used by Example GeneMapper ID Software v3 1 for a panel definition in a tab delimited text file txt using notepad or wordpad or a Microsoft Excel spreadsheet saved as a txt file hr AmpFLSTR_Panels_v 1 Notepad File Edit Format Help PGenemMapper ID v3 1 Last edited 071403 GMv3 0 For import into
338. urned on in the plot profile the word Changed displays inside the label box Allele Edit Comment Ctrl O Controls whether or not the Allele Comment dialog box opens when you make label changes Checked when active To view the comments entered e Show the allele edit AE comment column in the Project Window Genotypes Table or e Select the History option when right clicking a selected labeled peak or e Double click a label GeneMapper ID Software v3 1 User Guide Plot Window Menus Alleles menu items and descriptions continued Item Description Add Bin Ctrl B Allows you to add a marker bin as follows 1 Select the Binning mode 2 Click the Marker Range for the bin A line and a cross hair opens on the electropherogram a Select the Add Bin menu item or right click to open the Add Bin option b Add a bin by either dropping the cross hair at the center of the desired bin location or by dragging across the desired bin range The Edit Bin dialog box opens allowing you to add bin information Add Bin to D651574 x Name in 156 38 Location Left offset joao New offset default Right offsetlo 40 New offset default l Mutant Bin OK Cancel Edit Bin Ctrl U Allows you to edit a marker bin as follows 1 Select Binning mode 2 Select the bin to be edited 3 Select the Edit Bin menu item or right click the selected bin to o
339. us range Resize columns Click and drag between columns to change column width Quick sort column Shift click the column header to sort ascending shift click again to sort descending Deselect Row Ctrl click the Row Header box C 6 GeneMapper ID Software v3 1 User Guide Genotypes View Overview of the Project Window The Genotypes view shown below displays the results of allele calling analysis Each row or record in the table is a single marker for a sample For example if a panel specified for a sample contains four markers the Genotypes table contains four records for that sample each in a separate row F GeneMapper ID 3 1 Database gmid Is Logged In File Edit Analysis view Tools Help 18 x ek gt PS L OO EW f OR Table setina amor LsTR Table EENES B EPaneis Samples Genotypes Efjidertifier_v Sample Name Run Name Panel Marker Dye Allele1 Alee2 ADO AE os BiN PHR LPH SPU an BD CC ovl ce 19 Sample5 Databasing identtifler_v1 AMEL R KO Y BEEBEE EEG m 20 Sample3 Databasing Identifiler_v1 CSF1PO B 10 12 a C C C C E E a 21 Sample4 Databasing Identifiler_v1 CSF1PO B 10 13 C E E C a E E fEl 22 Sample3 Databasing Identifiler_v1 0138317 G 9 13 a C ia C C E E E 23 Sample4 Databasing identifie
340. user installation of software or interfacing or use in combination with software or products not supplied or authorized by Applied Biosystems and modification or repair of the product not authorized by Applied Biosystems The foregoing provisions set forth Applied Biosystems sole and exclusive representations warranties and obligations with respect to its products and Applied Biosystems makes no other warranty of any kind whatsoever expressed or implied including without limitation warranties of merchantability and fitness for a particular purpose whether arising from a statute or otherwise in law or from a course of dealing or usage of trade all of which are expressly disclaimed Warranty The remedies provided herein are the buyer s sole and exclusive Limitations remedies WITHOUT LIMITING THE GENERALITY OF THE FOREGOING IN NO EVENT SHALL APPLIED BIOSYSTEMS BE LIABLE WHETHER IN CONTRACT TORT WARRANTY OR UNDER ANY STATUTE INCLUDING WITHOUT LIMITATION ANY TRADE PRACTICE UNFAIR COMPETITION OR OTHER STATUTE OF SIMILAR IMPORT OR ON ANY OTHER BASIS FOR DIRECT INDIRECT PUNITIVE INCIDENTAL MULTIPLE CONSEQUENTIAL OR SPECIAL DAMAGES SUSTAINED BY THE BUYER OR ANY OTHER PERSON OR ENTITY WHETHER OR NOT FORESEEABLE AND WHETHER OR NOT APPLIED BIOSYSTEMS IS ADVISED OF THE POSSIBILITY OF SUCH DAMAGES INCLUDING WITHOUT LIMITATION DAMAGES ARISING FROM OR RELATED TO LOSS OF USE LOSS OF GeneMapper ID Software v3 1 User Guide J
341. ut off value Global cut off value verification using Classic and Advanced algorithms were evaluated using the defined ABI PRISM platforms and AmpF STR kits The cut off value ignores peaks lower than the cut off ratio as defined of the highest peak in the locus size range Three different values were chosen to test 60 alleles for the global cut off values of 0 10 and 20 The results show that the algorithms implemented in GeneMapper ID Software v3 1 properly filter peaks that meet the global cut off specification In Figure I 15 the higher the cut off value the less conservative the call meaning less peaks are detected Note GeneMapper ID software users should carefully consider the sample type when choosing the cut off value l 24 GeneMapper ID Software v3 1 User Guide E Genotypes Plot File Edit View Tools Alleles Help Results of Verification Testing Plot Setting HID_Genot notype i a ee Sample Name Pand z E Panes fs EP Ea lL 0 Cut Off Identifile_vl1 10 Cut Off l cro g mm A m me em im i me A CSFIPO 37_22_Itod Identifiler_v1 20 Cut Off carci e im a e ie ie ie CSFIPO Minus A Cut off Value Testing Figure l 15 Example of a1 to 9 DNA mixture sample tested using three global cut off values The second parameter tested in the Algorithm testing category is t
342. vide accurate base calling The GeneScan 500 250 size standard contains all of the fragments in the GeneScan GS 500 Size Standard except for the 250 bp fragment Since it is identical in other respects it is not discussed separately in this section How It Is The GeneScan 500 all Size Standard is prepared by PstI digestion of Prepared plasmid DNA followed by ligation of a LIZ or ROX dye labeled 22 mer oligodeoxynucleotide to the cut ends A subsequent enzymatic digestion with BstUI yields DNA fragments containing a single ROX or LIZ dye see GeneScan 500 Molecular Lengths below GeneScan 500 The following table lists the GeneScan 500 denatured fragment Molecular molecular lengths nucleotides for the 16 fragments Lengths Fragment Fragment Fragment Length Length Length 35 160 400 50 200 450 75 250 490 100 300 500 139 340 150 350 GeneMapper ID Software v3 1 User Guide D 7 Appendix D GeneScan Size Standards Electrophero The screen below shows an electropherogram of GeneScan 500 size gram of standard run under denaturing conditions GeneScan 500 Size Match Editor Edit View Size Match Quality 0 98 439 150160 dA tt L 6176 6492 6808 7124 7440 7756 8072 8388 8704 D 8 GeneMapper ID Software v3 1 User Guide Converting Macintosh Sample Files This appendix describes how to use sample file conv
343. ware e Only one user can access a database at a time After you enter the registration code see Registering GeneMapper ID Software on page 2 6 the Login screen appears You must log on to the GeneMapper ID software program with a user name so that the system can log information to the database and preserve the options for each user The first time you log on to the GeneMapper JD software the following dialog box opens F Login to GeneMapper ID 3 1 User Mame farid bd Fassword Database Host farsan z 1 _ 44 it j i A 4 j Applied Biosystems 1999 2003 Applera Corp All Rights Reserved Mew Host Delete Host To create a password 1 The first time you log on leave the User Name as gmid and leave the Password field blank 2 Select the Database Host local computer from the drop down menu and click OK The software opens a dialog box to change or create your password GeneMapper ID Software v3 1 User Guide 2 7 Chapter 2 Getting Started with GeneMapper ID Software 3 Create a new password Note The first time you use the GeneMapper JD software leave the User Name as gmid and the Old Password field blank a Enter anew User defined password b Confirm the User defined password and click OK 4 For subsequent launches of the GeneMapper ID software the following screen appears F Login to GeneMapper ID 3 1 User Mame fami bi Password Database Host farsan bi
344. ware v3 1 Generic Updater GeneMapper ID Software v3 1 Demo to Final Upgrade Software Utilities Apple Mac Conversion Utilities Figure 2 1 Installation options on the GeneMapper ID software installation CD ROM 2 4 GeneMapper ID Software v3 1 User Guide Installing the GeneMapper ID Software What gets The installer on the CD installs Installed e Oracle database GeneMapper I D software version e Panel folder that contains AmpF STR panels and bins e Microsatellite and SNP genotyping example data e Electronic PDF version of the user guide e Size standard folder which contains various size standards e Default plot and table settings GeneMapper GeneMapper Select an item bo view iks description See alsa My Documents My Network Places My Computer app Automation Contig Database J docs Example Data oMa 1 Database Defaults installer JRE Panels Plot Settings Size Standards Table Settings componentinstall GeneMapper_Log GeneMapper Preferences a GeneMapper_ Resources install Service License Properties Pnr E uninstall Figure 2 2 Example of the installed GeneMapper folder GeneMapper ID Software v3 1 User Guide 2 5 Chapter 2 Getting Started with GeneMapper ID Software Registering GeneMapper D Software Completing the Registration Card Registering GeneMapper D Software Complete the registration card included in this package and return it t
345. ways reads GeneMapper HID Allele Calling Algorithm This action is displayed when you add a label manually to a non labeled peak Deleted when you remove a label from a labeled peak Edited when you change the label of a labeled peak Note If you make multiple edits to an allele then delete the label All of the previous edits are NOT listed in the history Edits are only shown for peaks that have labels Similarly allele edit comments are only shown in the genotypes table for peaks that have labels 7 26 GeneMapper ID Software v3 1 User Guide Genotypes Table Genotypes Table IMPORTANT If the overall size quality PQV displayed for all samples is red low quality the genotypes table are not accessible After viewing plots and editing alleles as necessary you can view the genotypes genotype PQVs and the overall Genotype Quality in the Genotypes table This feature provides a tabular format to display the genotypes of your samples and the quality of the samples Each row corresponds to a locus within a sample Therefore if you have 9 loci per sample you will have 9 rows for that sample within the genotypes tab This display is also known as a vertical table format The software is not capable of producing a horizontal table format where each row corresponds to a single sample To create a horizontal table export the genotypes table and make changes using a downstream spreadsheet application such as Mi
346. work 96261 2002 08 01 15 33 47M Bo E Datahace Praiart anns 7 4 Ad AN hd pen Cancel Note Once you have opened a Project window you may resize it horizontally and vertically to increase the width and depth of the Project table The following table describes the Open Project dialog box window elements Element Description Search field Typing in this field selects the first item in the project list whose name matches the characters GeneMapper ID Software v3 1 User Guide Project Window File Menu Dialog Boxes Element Description Project table Lists all the projects in the database e Project Name of the project e Last Saved Date and time stamp of when the project was last modified e Owner Name of the user who created the project e of Samples the number of samples in the project Note Columns may not be moved or hidden You may resize the width of the columns by dragging between the column headers Only one row in the table may be selected at a time You may navigate up and down in the table using arrow keys Shift clicking a column header sorts the data by that column Open Opens the selected project If previous project has pending changes the following alert message is displayed before the Open Project dialog box opens Do you want to save changes Yes No Cancel Cancel Closes the Project Manager dialog box Save Project and The Save
347. x A Process Quality Values explains how to use the Process Component Based Quality Values e Appendix B Software Genotyping Algorithms provides a description of the genotyping algorithms used in GeneMapper ID software e Appendix C Project Window Software Interface describes the dialog boxes and menu commands associated with the Project Window e Appendix D GeneScan Size Standards provides information on the defined size standards provided with the GeneMapper ID software e Appendix E Converting Macintosh Sample Files describes how to use the Macintosh AppleScript program to convert Macintosh generated fragment analysis sample files to the Windows format e Appendix F Transferring Data and Backing Up Data describes how to transfer projects supporting files and data to a different computer running GeneMapper JD software e Appendix G Sample Sheets and Plate Records e Appendix H Troubleshooting Sheet e Appendix I GeneMapper ID Software Verification describes verification testing e Appendix J Software Warranty describes the Applied Biosystems warranty that comes with the software package e Appendix K Workflow for Storing Sample Files Projects and Second Reads GeneMapper ID Software v3 1 User Guide 1 7 Chapter 1 About GeneMapper ID Software 1 8 GeneMapper ID Software v3 1 User Guide Getting Started with GeneMapper D Softw
348. xport the file b In the Export File As section select CMF 3 0 xml from the drop down list Note GeneMapper ID software supports CMF 1 0 dat file types for export c In the CODIS Laboratory IDs section select a source lab from the Source drop down list and a destination lab from the Destination drop down list d Type in the desired File Name The default file name is the name of the project e Click Export GeneMapper ID Software v3 1 User Guide 9 9 Chapter 9 Exporting Table Data GeneMapper ID Software v3 1 User Guide Printing Data This chapter covers PUIG DACA cts aerirare a aS ohana 4 etal ele idty st 10 2 Pace Ser p OPO esas oot ow ete SS ea the aes ee aedes 10 4 Printing Sizing Plots Similar to Genescan Using the Predefined AmpF STR Sizing Plot Setting 10 7 Printing Genotype Plots Similar to Genotyper Using the Predefined AmpF STR Genotyping Setting 10 8 GeneMapper ID Software v3 1 User Guide 10 1 Chapter 10 Printing Data Printing Data Printable Views In the GeneMapper ID software you can print the following views e Samples table e Genotypes table e Samples plots e Genotypes plots e Raw data e Sample info e EPT data To print one of the views listed above display the view on your screen then select File gt Print For example if you want to print the raw data of a particular sample select the sample in the Samples tab then se
349. y LIZ to check sizing precision when using a LIZ size standard 4 ik Samples Plot File Edit view Tools Alleles Help Plot Setting Overlay LIZ Dye v E Panes fi zji eee E a Ei kal l ee majem Pi EES LA PS Sample Name Pand sgo SQ here are no controls in this selection 100 200 300 400 500 1200 1000 800 600 400 200 Dye Sample Peak Sample File Name Marker Allele T i ID_Sample3 fsa ID_Sample4 fsa ID_Samples tsa X 218 53 Y 962 GeneMapper ID Software v3 1 User Guide 7 21 Chapter 7 Analyzing and Interpreting Data Overlay ROX Use Overlay ROX to check sizing precision when using a ROX size standard Samples Plot Ioj xj Fie Edit view Tools Alleles Help Plot Setting Overlay ROX Dye Panes fi oe al Emha lei majen i af ia w Sample Name Qo0 SQ ee are no controls in this selection 120 130 150 4160 170 180 130 200 210a e20 2350 240 250 260 270 280 230 300 31410 320 2000 1800 1600 1400 1200 1000 300 600 400 200 0 n Dye Sample Peak veal File Name Marker Allele PP_Sample2 fsa xX 120 43 Y 1109 These factory provided defaults should be used as a guideline to customize your plots Refer to Chapter 5 Section 5 4 for information on creating and saving plot settings and refer to Chapter 8 for detailed information about plot window toolbars and menus Once you have created custo
350. y indications A 5 Index 6 SSCP Minimum Peak Half width setting B 10 B 15 startup options 6 3 Stutter Distance 5 18 Stutter Ratio 5 18 stutter ratio 7 3 J table filter using 8 19 Table Manager content column dialog box 5 33 filtering controls 5 35 Table Setting Editor copying table profiles 5 34 deleting a table profile 5 35 editing a table profile 5 34 filtering controls 5 35 filtering settings genotypes view 5 37 filtering settings samples view 5 36 new table profiles 5 34 Table Settings Editor 5 29 General tab 5 30 Genotypes tab 5 31 Sample and Genotypes tab elements 5 32 Sample and Genotypes tab elements allele settings 5 33 Samples tab 5 31 Table Settings tab profiles 5 28 5 42 Technical Communications contacting xi e mail address xi Technical support for computers with altered configuration J 2 Technical Support contacting xi Thresholds 5 24 Training obtaining information about xi transferring data F 2 troubleshooting bad matrix files causes 5 64 GeneMapper ID Software v3 1 User Guide U UDI for CODIS Specimen Number 9 8 for shared names 9 8 User attention words defined x User guide organization 1 6 W Warranty exceptions J 3 for computers with altered configuration J 2 Warranty limitations J 3 workflow for storing sample files projects and second reads K 1 GeneMapper ID Software v3 1 User Guide Index 7 Index 8 GeneMapper ID Software v3 1 User Guide
351. y or Click Drag inside the plot area e Select discontinuous peak selection Use the CTRL key The selected peak is maintained when switching between the Samples and Genotypes tabs in the Project window with the Plot view still visible GeneMapper ID Software v3 1 User Guide 8 23 Chapter 8 Using Plot Windows Samples and Genotypes Alleles menu items and descriptions continued Item Description Editing Mode continued In Binning mode you can e Edit marker size ranges e Edit bins When the Binning mode is selected the Plot window automatically switches the display configuration to an appropriate view for editing bins Plots shown in Separate Dyes mode for microsatellite samples Plots shown in Combine Dyes mode for SNP genotyping samples X axis displayed in base pairs Marker Indicator and Marker Range displayed Bins displayed To edit marker size ranges for microsatellites only 1 Select Binning mode 2 Click the marker indicator of the marker to be edited A red indicator displays 3 Drag the red indicator line to the desired size range Note To edit allele calls refer to Add Allele Delete Allele and Rename Allele rows in this table To edit Bins refer to Add Bin Edit Bin and Delete Bin rows in this table To edit marker size ranges for SNPs the bins need to be edited Binning Mode is disabled when e There is no bin set for the panel e The
352. y_input Profiler_plus_NT Profiler_Plus_v1 FGA B ial E E A B E E E A lovy_input Profiler_plus_NT Profiler_Plus_v1 VNA B w f A w E B w A lovy_input Profiler_plus_NT Profiler_Plus_v1 AMEL G a m m E w m E lovy_input Profiler_plus_NT Profiler_Plus_v1 D13S317 Y E A a a El iz A lovy_input Profiler_plus_NT Profiler_Plus_v1 D18S51 G a A a A m E B o low_input Profiler_plus_NT Profiler_Plus_v1 D21511 G E E A A E E E E A lovy_input Profiler_plus_NT Profiler_Plus_v1 D3S1358 B E a E B E E E A q low _input Profiler_plus_NT Profiler_Plus_v1 D7S820 E A A E E E B A i TE ow ipat PrO atest 5 49 lovy_input Profiler_plus_NT Profiler_Plus_v1 FGA B 20 22 E w w m bei m 50 low _input Profiler _plus_NT Profiler_Plus_v1 VWA B 16 18 E E E E E a A 51 lovy_input Profiler_plus_NT Profiler_Plus_v1 AMEL G Xx M M E E Ei o E 52 low input Profiler_plus_NT Profiler_Plus_v1 D13S317 Y 10 11 E A A E E E E 53 lovy_input Profiler_plus_NT Profiler_Plus_v1 D18551 G i E E Ei E B E mE E 54 low _input Profiler_plus_NT Profiler_Plus_v1 D21511 G 242 OL ii A A E E E 55 lovy_input Profiler_plus_NT Profiler_Plus_v1 D3S1358 B 14 17 cal al E w w E B E 56 lovy_input Profiler_plus_NT Profiler_Plus_v1 D55818 Y J11 Bw E E i a E 57 low input Profiler_plus_NT Profiler_Plus_v1 D75820 Y 13 E E E E E a A 58 low_input Profiler_plus_NT Profiler_Plus_v1 D8S1179 G 12 16 E E E A E E E E 59
353. yping e AmpF STR Sizing Last Used Plot Overlay LIZ Overlay ROX You need to import these default plots into the database through the Plot Settings tab To import a default plot setting l 2 Press Import A dialog box opens Double click the Plot Settings folder to display all of the default plots Select the default plot you wish to import then click Import The default plot appears in the Plot Settings tab GeneMapper ID Software v3 1 User Guide Section 5 4 Plot Settings Tab Plot Settings Editor Introduction How to Access the Plot Settings Editor General Tab The Plot Settings Editor allows you to customize your plot views by adjusting the following e The items displayed in the Sample headers e The items displayed in the Genotype headers e The columns displayed in the Sizing table e How labels are shown e Display settings The Plot Settings Editor can be accessed in the following ways e Select Tools gt GeneMapper Manager gt Plot Settings Tab gt New or Open e Inthe Sample or Genotype Plot views select a profile to edit and select Tools gt Plot Settings or click the Plot Setting Editor icon on the toolbar The General tab allows you to enter in the name and description of the plot setting Plot Settings Editor x General Sample Header Genotype Header Sizing Table Labels Display Settings When Opening The Plotyyvindo Mame New Plot Setting o Can

draft - Thermo Fisher Scientific

Contents

Download Pdf Manuals

Related Search

Related Contents