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Applied Biosystems Prism 377 GeneMapper Software User Manual

1. Microsatellite E Panes 2 z TE Gee Bo walt EAH a MA ie Malae orara eS Sample File Sample Name Panel 0S SQ g 39 793 M9 459 499 239 279 39 359 399 439 479 519 559 599 633 673 719 4 8398 7464 6531 5598 3732 2799 18664 u ri Plots The Overlay All option can be used to verify size standards continued as follows 1 Select any number of Samples in the Project window 2 Click tl Display Plots 3 Turn off all dye color icons except the dye color used for the size standard 4 Click E Overlay All All size standard electropherograms are displayed this allows the user to check for outliers Using Plot Windows Samples and Genotypes 7 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Tables Controls which table if any is viewed at the bottom of the Sample Plot window No Table Ctrl Q Table Filter gt Sizing Table Ctri A v Header Genotypes Table Ctrl V No Table Ctrl Q Table is turned off Sizing Table Ctrl A A sizing table is shown 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 Genotypes Table Ctrl W The ge
2. 1 4 About GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Unique features of the GeneMapper software v3 0 continued Unique Feature Description GeneMapper Database The GeneMapper database stores the following data Predefined size standard definitions included with installation and custom size standard definitions Panel marker and allele bin definitions Analysis methods Table profiles for generating tabular reports gt gt OH Projects containing analyzed data allele calls confidence values analysis methods and size standard Automated operation for all but Automated allele calling is made possible by instrument and problematic data software calibration and setup steps done once for many samples including user defined criteria It is also made possible by the PQV system described above Bad or out of bounds data can be safely discarded and only data with minor component problems is deferred for user intervention About GeneMapper Software 1 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Important Features GeneMapper software v3 0 has a number of features that are built on the automated genotyping capabilities of the ABI PRISM GeneScan and Genotyper applications The table below lists important features of the GeneMapper s
3. Plot Properties Control Print dialog box 3 2 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Windows and Utilities Supporting the GeneMapper Software Project The figure on page 3 2 shows the Project window as the main window and the manager utilities editors and dialog boxes as the subordinate windows These windows are opened from and provide support for the project As the focal point of the GeneMapper application the Project window makes it easy for you to Add samples initiate analysis and export results View sample data and access data plots You can view the data associated with a particular sample file by Viewing the pre analysis raw data version of the data in the Samples view of the Project window Viewing the tabular results of analysis in the Genotypes view of the Project window or Viewing plot s of selected Project sample data in the Plot window Initiate the process of creating analysis methods and defining panels and bins The following items support the information in the Project window and are available in the GeneMapper Manager in the Project window Table Setting Editor GeneMapper Manager This page enables you to change the way information is displayed in the Project window including Hide Show table columns in the Project window Filter table entries Set pro
4. E Project Info Raw Data EPT Data 4 SNaPshot 2 5 Linkage 01_100301 fsa 4400 1838 2276 2714 3152 3590 4028 4466 4904 5342 5780 6218 6656 7094 02_1004 319F g i 03_1005 oral 04_10031 05 1004 a 06_1005 w 07_1003 135 s 43 3 EP Voltage m EP Current pa EP Power mmm RunTemp pE Line Description Blue EP Voltage Black EP Power Green EP Current Red Run Temperature 3 10 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Samples and Genotypes Views There are several plot sealing features in the EPT view 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 Return to a full scale plot by double clicking in the label area again The plot window is restored to full scale You can scale the Y axis to maximum Y or a user defined value by selecting the Y Axis Scale from the View menu or specify upper and lower limits in the Scale to dialog box The Samples and Genotypes tabs provide access to two separate tables each of which is considered a different view of the same project data Only one of these views is displayed in the Project window at any time Samples View See page 3 12 and in detail on page C 2 This view is used to enter samples and its tab is always enab
5. Show Records Containing Show All Records Show All Records Show Records Containing Note This setting allows specification by a string Input this information into the Content column Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotypes view filter settings continued Column Name Filter Settings Dye Blue Green Orange Red or Yellow 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 SNaPshot data cannot be filtered by this selection Off scale Sharp Peak M One Basepair Allele M Single Peak Artifact M Split Peak M Out of Bin Allele Peak Height Ratio 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 Records Show All Records Using Table Settings 6 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotypes view filter settings continued Column Name Filter Settings Genotype Quality GQ Show Pass Show Check or Show Low Quality Show All Re
6. The Project window enables you to Minimize the window to an icon on the Microsoft Windows taskbar or to the title bar Maximize the window to fill the entire screen Resize the window by dragging any window edge or corner The following reformatting can be done in the Project window to change the presentation of information The View menu provides a Show Navigator toggle command to show hide the navigation pane for both the Samples and Genotypes views When the navigation pane is hidden the table occupies the entire width of the Project window Shift clicking a column header sorts 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 47 The first column of the table displaying row numbers is fixed during horizontal scrolling of the Samples and Genotypes tables Using the Project Window 3 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com You can create Hide Show columns as needed with different table settings created using the Table Settings in the GeneMapper Manager window Resize columns by dragging the separating lines When resizing is enabled the cursor changes to a resize cursor a double headed arrow that looks like this lt Parts of the Main Once you have loaded Sample files into a Project window the main Window window looks like the example
7. GeneMapper Help F1 About GeneMapper Project Window Help Menu Commands Item Description Enabling GeneMapper Help Opens the GeneMapper User Always enabled F1 Manual PDF file using Acrobat Reader Does not open the tutorial guides About Opens the About GeneMapper Always enabled GeneMapper window and displays the software version C 26 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 36 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 amp p Save Project x 4 i Project name Cancel Project Window Software Interface C 27 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Open Project The Open Project dialog box displays the list of projects in the Dialog Box database You use the first field to find a p
8. Open the Clear the And GeneMapper software displays the Navigation pane View menu from the Project window Alternative Ctrl Shift N Show Project window without the navigation pane j Navigator File Edit Analysis View Help check mark ce ees Satie otic smena Samples Genotypes Status Sample Name Sample Type Analysis Method Panel Size Standard Run Name Run Date amp l E Jio Sample demo_LMS Panel01 M ManGS400HD Linkage 1998 02 27 z 100405 Sample demo_LMS Panel01 MManGS400HD Linkage 1998 02 27 3 E r000 Sample demo_LMS _ Panel01 ManGS400HD Linkage 1998 02 27 4 100302 Sample demo_LMS _ Panel01 t ManGS400HD Linkage 1998 02 27 s 100405 Sample demo LMS Panel01 t ManGS400HD Linkage 1998 02 27 e m 100502 Sample demo_LMS Panel01 MManG5400HD Linkage 1998 02 27 7 100305 Sample demo_LMS Panel01 t ManGS400HD Linkage 1998 02 27 To see the Click the And GeneMapper software displays the Genotypes view Genotypes tab Project in the Genotypes view GeneMapper Project LMSdemo Alternatives Select View gt Samples or Ctrl Shift 1 File Edit Analysis View Help Alternatives File Edit Analysis View Help SOW MW M Tabieprone fapoicasiosystemsveraut Flim BS Select S S Panels Samples Genotypes Genotypes Baleara R
9. Project column Project name Last Saved column Date Time stamp showing when the project was last saved Owner column User name of the person who created the project 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 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 3 24 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Export Project The Export Project window enables you to export a project out of the Window GeneMapper database into a specified location See an explanation of each area on this window in Using the Export Project Window on page 3 26 1 2 E Export Project 3 4 5
10. Click the Raw Data tab What to Look For In the raw data display of the sample files verify the following in the Raw Data Data peaks are present in all four or five of the matrix standards Display gt There are no anomalies The baseline is stable flat between peaks Peaks are on scale no more than 8191 relative fluorescent units and the peaks of the dye of interest have a value of at least 200 9 10 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com If peak data does not show these characteristics refer to Causes for Bad Matrices on page 9 19 for possible interpretations of your peak data Choosing a Data To choose a data point range Point Range Step Action 1 Move the cursor well away from the primer peak in a region at the 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 You will need to enter these values in the next step when generating the new matrix refer to page 9 12 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 dis
11. If Allele Edit Comment is selected the Add Allele Comment dialog box opens allowing the user 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 the user renames 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 Allows the user to delete an allele call from a labeled peak Delete as follows a Select Peak Selection mode b Select a labeled peak using the left mouse button c 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 the user to enter an allele comment for the 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 the user to view the allele history in columns labeled Basepair All
12. Y Axis Scale Controls the vertical scale of the electropherograms currently viewed as follows Scale Individually Each electropherogram being viewed is scaled to its maximum height Scale To Maximum Y All plots currently viewed are scaled to a global maximum value 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 gt Scale To Maximum Y v Bins Ctrl B Scale To Zooms in on a specific region on the Y axis of the electropherogram as follows Click and drag with the left mouse button the desired region on the Y axis This only affects the selected electropherogram or 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 ty Y Axis Zooming x M Y Axis zoom To Tick Spacing unitsitick I Apply to all electropherogram Cancel Double click on the Y Axis to return selected electropherograms to Full View Shift double click on the Y Axis to return all electropherograms to Full View Using Plot Windows Samples and Genotypes 7 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Labels Controls ho
13. A rtisan Artisan Technology Group is your source for quality TecmoogyGroup new and certified used pre owned equipment FAST SHIPPING AND SERVICE CENTER REPAIRS WE BUY USED EQUIPMENT DELIVERY Experienced engineers and technicians on staff Sell your excess underutilized and idle used equipment TENS OF THOUSANDS OF at our full service in house repair center We also offer credit for buy backs and trade ins IN STOCK ITEMS www artisantg com WeBuyEquipment 7 EQUIPMENT DEMOS HUNDREDS OF InstraV ea REMOTE INSPECTION LOOKING FOR MORE INFORMATION MANUFACTURERS Remotely inspect equipment before purchasing with Visit us on the web at www artisantg com 7 for more our interactive website at www instraview com information on price quotations drivers technical LEASING MONTHLY specifications manuals and documentation RENTALS ITAR CERTIFIED D a gaa tia Contact us 888 88 SOURCE sales artisantg com www artisantg com ABI PRism GeneMapper Software Version 3 0 User s Manual gt Applied KS Biosystems Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Copyright 2002 Applied Biosystems All rights reserved For Research Use Only Not for use in diagnostic procedures 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 b
14. Note For information on the tick marks displayed in the electropherogram see Examining Peak Definitions on page B 19 Software Genotyping Algorithms B 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Effects of The figure below shows that both alleles are detected after re analyzing Decreasing the with the polynomial degree set to 3 while decreasing the window size Window Size Value Value to 15 from 19 data points A 40 sso awo so seo soo 4200 42104220 4230 4240 4250 4200 4270 4280 4200 400 4310 4920 4300 4340 4960 400 4970 48O 400 4400 4410 4420 206 30 a e e OU OA 206 31 15 92 186 78 1983 42443 4342 Electropherogram showing the alleles detected as two peaks after decreasing the window size value B 24 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Optimizing Peak Detection Sensitivity Example 2 Initial The figure below shows an analysis performed using a polynomial Electropherogram degree of 3 and a peak window size of 19 data points AJ 1820 1890 1840 1860 1860 1870 1880 1890 1900 1910 1920 1990 1940 1960 1900 1970 1980 1990 2000 2010 2020 2090 2040 La y Dye Sample Minutes Size Peak Height Peak Area Data Point Peak BR 15 5 18 339 50 1402 _ 17703 1935 3R 16 6 19 341
15. Plots Controls the way electropherograms of samples are shown in the Sample Plot window Tables gt Separate Dyes Ctrl 8 Table Filter gt Overlay All Ctri 9 Combine Dyes All dye colors from a sample are combined into a single source electropherogram penai Jm me E EE EEE Ma ew MaE Sample File Sample Name Pand os sQ 14 43 15 17 19 24 23 25 27 29 3 33 35 37 39 4 43 45 47 49 51 53 55 57 59 61 63 65 67 69 7m 73 3824 7 18 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Plots Separate Dyes Each dye color from a sample is continued separated into an individual electropherogram Microsatente K Panes 2 Ge Ge Bh uw Has i el Tae s Sample File Sample Name Pand os sQ m no 4 pais isas IORI CERI CEEI 68 86 106 126 148 166 136 206 226 24 266 286 306 32 346 36 38 406 maa mp pan maf al 336a 3 aocsen tex 100501 Seeo1 pana o1 E no 4 66 86 108 126 146 166 186 206 226 24 266 28 306 325 34 36 36 406 fal 17652 Overlay All Multiple samples are combined into a single electropherogram
16. Software Version 3 0 PN 4335524 Portions of this chapter are organized around the Panel Manager menu items to explain various features and actions that can be performed In This Chapter This chapter contains the following topics Topic See Page Panel Manager Overview 4 2 Panel Manager Window Commands 4 6 Formats of Panel and Bin Text Files 4 23 Panel Table View 4 33 Marker Table View 4 35 Bin View for Microsatellites Only 4 37 Using Panel Manager 4 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Manager Overview Introduction The Panel Manager utility allows you to manage all of the chemistry kits panels markers and bin definitions required for automated allele calling Topic See Page Purpose of the Panel Manager Window 4 2 Displaying the Panel Manager Window 4 3 Hierarchy of Panel Data 4 3 Panel Manager Toolbar 4 4 Panel Manager Navigation Pane 4 5 There are two ways to get data for the Panel Manager Creating kits panels markers and bins manually Importing preformatted text files that contain panels markers and bin definitions See Formats of Panel and Bin Text Files on page 4 23 for two examples Purpose of the The Panel Manager window enables you to create edit and import Panel Manager panels It features a Window Tool for creating kit folders Table for spe
17. The following table lists how to access subordinate GeneMapper Access to Subordinate software windows Application Windows Access GeneMapper Application Windows To see the Select And GeneMapper software displays the Analysis Tools gt Analysis Methods page Methods GeneMapper Manager zi Table Settings Plot Settings Matrices Size Standards Then select Last Saved Owner Instrument Analysis Type Description the Analysis 1 3100Avant DS33 2002 08 20 13 5 GM Microsatellite 2 3100Avant SNP 2002 08 20 13 5 GM 8NaPshot Methods tab 3 3730 DS 33 Install 2002 08 20 13 0 gm 3730 Microsatellite Factory Provided 4 Default 2002 08 20 13 0 6M Microsatellite 5 Microsatellite Default 2002 08 20 13 0 gm Microsatellite Factory Provided 6 SNaPshot Default 2002 08 20 13 0 gm SNaPshot Factory Provided Delete Done Panel Manager Tools gt Panel Manager or click the AE aii EG a icon EK a EEE mstr E Panel Manager Pavel Name Comment E Bues Jrone Green_l_v3 none Profiler_v3 none Profiler_Plus_v3 none ee COfler v3 none SGM_Plus_v3 none Identifiler_v3 none Profiler _Plus_CODIS_ none COfiler_CODIS_v3 none O Identifiler_CODIS_v3 none Panel Manager window OK Cancel Apply Artisan Technology Group Quality Instrumentation Using the Project Window 3 15 Guarante
18. 39 79 493 459 499 239 279 39 359 399 439 479 519 559 599 633 679 M9 A 7733 68734 6014 siss Full View X Axes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Plot Windows Samples and Genotypes 7 9 Item Description X Axis Scale Selects the horizontal scale units for the X axis to be displayed in either base pairs default or data points X cale Basepairs Ctri Y Y Ais Scale gt Data Point Ctri N Zooms in on a specific region on the X axis of the electropherogram as follows Click and drag with the left mouse button the desired region on the X axis In the Samples Plot this affects all electropherograms In the Genotypes Plot this affects only the selected electropherogram or 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 Gp X Axis Zooming s x X AX S Zoom From to Tick Spacing units tick Cancel 7 10 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description
19. A graphical editor for defining bins For more details see Appendix C Project Window Software Interface Size Standard Editor GeneMapper Manager This window enables you to create new size standard definitions to be used with new groups of samples and delete existing size standards For more details see Creating Editing a Size Standard on page 10 5 Size Match Editor This window enables you to adjust an in lane size standard to compensate for peak shift or a missing peak For more details see Size Match Editor on page 10 12 Display Plots This window enables you to display graphically the data associated with samples and markers and visually assess the quality of the data For more details see Displaying Plot Windows on page 7 2 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Details of the GeneMapper Project Window Introduction The following information is provided for the GeneMapper Project Project Window Interface Conventions Reformatting the Window window Topic See Page Project Window Interface Conventions 3 5 Reformatting the Window 3 5 Parts of the Main Window 3 6 Navigating in the Project Window 3 7 Samples and Genotypes Views 3 11 Project Window Toolbar 3 12 Viewing the Project Window Contents 3 14 Access to Subordinate Application Windows 3 15 Column Menus and Rules for Columns 3 18
20. Creating and Evaluating a Matrix describes how to create a matrix file Chapter 10 Using the Size Standard Settings describes how to create a size standard and how to use the Size Match Editor The following appendices contain reference information 2 12 Using GeneMapper Software Appendix A Process Quality Values explains how to use the Process Component Based Quality Values Appendix B Software Genotyping Algorithms provides a description of the genotyping algorithms used in GeneMapper software Appendix C Project Window Software Interface describes the dialog boxes and menu commands associated with the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Appendix D GeneScan Size Standards provides information on the Size Standards provided with the GeneMapper software Appendix E Sample File Conversion describes how to use the Macintosh AppleScript program to convert Macintosh generated fragment analysis sample files to the Windows format Appendix F Software Warranty Information describes the Applied Biosystems warranty that comes with the software package Using GeneMapper Software 2 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE
21. Note The Plot Window cannot contain electropherograms from both the Samples and Genotypes tabs at the same time Certain features within the Plot Windows can be customized using 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 See Chapter 8 Using the Plot Settings Editor The last profile used in the Samples Plot is used to open the Genotype plot The last profile used in the Genotypes Plot is used to open the Sample Plot Using Plot Windows Samples and Genotypes 7 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Plot Window Toolbars Toolbar The toolbar icons enable and disable plot features similar to their Descriptions associated menu items as described throughout this chapter Position your mouse over an icon to view its tooltip description 1 3 4 5 6 wierscateut EU Panes 2 EE Geil Br Mel Be fii 2 Sf TA jfa ii 7 8 9 10 Item 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 electropherogram panes shown 3 Dye color toggle icons View menu Header Marker Range and Marker Indicator toggle icons View menu 5 Icons t
22. Project dialog box again Cancel Closes the dialog box and does not save the project C 30 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Save Project Elements continued Element Description Save Saves the project under the name entered in the dialog box Don t Save Aborts the Save action Export Table The Export Table dialog box allows you to choose where to save the Dialog Box exported file The file types supported include Tab delimited text txt 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 2 ow Export Table x Lookin 23 DiGMdistributionExamples X E ck E aj PanelData SampleFiles 3 4 f 5 File name Export lable 6 Show files of type fall Files 7 Cancel 7 Export file as Tab delimited text bd z 8 Project Window Software Interface C 31 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Export Table Callouts Item Name Description 1 Toolbar These icons are used as follows Clicking moves display up one level in
23. bin name 1 GM version number 2 bin center 2 Kit name 3 bin left offset 3 bin set name 4 bin right offset 4 panel name 5 marker name Using Panel Manager 4 27 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Definitions for SNaPshot 4 28 Using Panel Manager The examples below show SNP panels defined in the format used by GeneMapper software v3 0 in a tab delimited text file txt or an Excel spreadsheet saved as a txt file fj SNPdemo txt Notepad loj x File Edit Format Help version GM v 3 0 Kit type SNP Chemistry Kit SNPdemo none Panel 6plexdemo none 20mer none 28mer none none none none none Columns 1 marker name 2 control allele names as comma separated bin names A G if none there must be a 3 comments if none write none Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Ed Microsoft Excel Book2 Mie xi E Eile Edit wiew Insert Format Tools Data window Help x GM 3 0 SNP SNPdemo none 6plexdermo none none none none none none Columns 1 marker name 2 control allele names as comma separated bin names A G example if none there must be a 3 comments if none write none Using Panel Manager 4 29 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www a
24. main pane Clicking moves display to Home level in main pane This is usually Profiles lt user gt cx Clicking creates a new folder at the present directory level ezez Clicking presents a list of the contents of the selected folder Clicking presents details of the selected folder 2 Drop down Select drive letter and or folder directory menu 3 Folder file The contents of the drive directory are selected display pane in the toolbar and the directory menu is displayed here 4 Export Table This button exports the selected table button 5 File name field Enter a file name to use for exported table 6 Show files of This is a display filter for files folders are always type field shown Allows specification of file extension Filters files for display in all files txt files csv files 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 export field Note Exported text tables show PQV s as numbers even if they are displayed as symbols C 32 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Software Interface 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 The dialog box
25. shows all files Java Serialized file ser shows only files with the extension ser which is how GeneMapper projects are exported Note There is only one preset file extension ser that can be used 7 Cancel button Closes the Export Project window without exporting a project 3 26 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 3 28 E Import Project 3 f 5 File name OO Import Files of type Java serializedfilecse I Cancel 6 7 Using the Project Window 3 27 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Import Project Window The elements called out in the figure in Import Project Window on page 3 27 are described in the table below Import Project Callouts Item Name Description 1 Toolbar These icons are used as follows Clicking moves display up one level in main pane Clicking moves display to Home level in main pane This is usually Profiles lt user gt cx Clicking creates a new folder at the present directory level B b 0m0 Bete Clicking presents a list
26. the appropriate size standard For SNaPshot kit reference data samples must have been analyzed using the appropriate size standard and an Analysis Method with SNaPshot as the Analysis Type Using Panel Manager 4 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Reference Data This feature now works for both microsatellite and SNP data SNP data The Panel SNP Reference Data command opens the SNP Reference Data dialog box allowing the user to organize the reference data imported at the kit level into panels This feature is only used when creating panels manually not when using the Auto Panel feature The panel is enabled when a SNaPshot panel is selected and reference data has already been added at the kit level ty SNP Reference Data SNP Kit reference data E x SNP Panel reference data 9 SNaPshot Tutorial 6plex Sample H A111347 15 fsa H A1Snapshot Control fsa A51347 01 fsa AT1347 02 fsa AI1347 101sa OK Cancel Microsatellite data The Panel Reference Data command allows you to remove reference samples from the selected microsatellite panel ty Microsatellite Reference Data x Microsatellite Panel reference data 29 DS 33 Remove OK Cancel 4 18 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Auto Bin The
27. 0 0 2 0 25 PlusA ratio 0 0 1 0 0 95 0 95 PlusA distance 0 0 Infinite 1 6 bp 1 6 bp Stutter ratio 0 0 1 0 0 95 0 15 Stutter distance 0 0 Infinite fromOto3 5bp from 0 to 4 5 bp Using Analysis Methods 5 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The parameters in the table except Cutoff value are defined graphically in the following figure Allele PlusA yy PlusA ratio B Stutter A Stutter f Stutter Distance PlusA Distance SNP Cutoff Value for SNaPshot For heterozygous SNP genotyping alleles if the highest peak height is X and a second lower peak height is Y then the ratio Y X must be larger than the cutoff value in order for the second peak to be called The default value is set at 0 3 and may need to be further optimized for your data Range Filter The Range Filter allows you to remove labels from peaks within a specific size range for each color 5 10 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Detector Tab Use the Peak Detector tab to select the algorithm to use for detecting peaks Basic Advanced Classic See Appendix B Software Genotyping Algorithms for additional information on Basic Advanced and Classic peak detection algorithms Basic Peak Detection Algorithm The default B
28. 440 4150 400 470 480 490 4200 4210 4220 4230 1000 Electropherogram showing the result of an analysis using extreme settings for peak detection Software Genotyping Algorithms B 27 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Parameters 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 on Baseline Q 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
29. 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 Minimum quality value 0 4 Allele Naming Scheme Number 1 2 3 4 C Letter A B C D C Rounded basepair 100 200 300 Auto Bin Options Auto Bin clear existing bins C Incremental Auto Bin keep existing bins Auto Panel The Auto Panel command opens the Auto Panel dialog box to automatically create 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 Ei Auto Panel xj Save Existing Panels Allele Names Maue 6 Yellow fo Micreen fA Wired Fo rMarker multiplex Minimum multiplex 5 Maximum multiplex po Panel Names Base Panel name rMarker Overlap Maximum overlap C No overlap Number of basepairs between markers p OK Cancel Using Panel Manager 4 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com View Menu Full View Ctrl x Axis Scale gt Y Axis Scale gt Dyes Full View Depending on the X and Y axes mode the Full View command resets the X or Y axis to the m
30. E CECI C CEEL In ee ha Na oo IRT 123 125 127 19 A 133 435 A7 49 AAA 443 145 447 449 A51 153 155 457 159 161 163 148 tt S es ee ee ee ee ee se ee ee ee SR SS 115 82 32 A aranan iN FEIR j J f 1 J 1 1 PE PRPS A Sets i INA fi 0 C06 a cus Setos Panel Dis2rHs B lg le ie ie EL a me jaa fe fo 278 160 162 464 166 168 230 4170 472 474 176 178 180 182 184 186 a a a A O M dart 488 190 192 194 19 198 1m aa oe ie le le le i ie fs fo ro 277 279 281 283 285 287 289 291 293 ag 295 gm Page 1 of 1 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 printed on the whole height of the page on a printer Project Window Software Interface C 35 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Add Samples to Project Dialog Introduction This section provides general information about the Add Samples to Project dialog the Edit and View menus available in the window using the three tabs in the window This following topics are covered in this section Topic See Page General Information and Procedure C 36 Edit Menu C 37 View Menu C 39 Files Tab C 40 GM Database Tab C 41 Sequence Collector Tab C 44
31. Enabled when analyzed samples are present in the table Non concordant Samples to Top Brings non concordant samples to the top of the analysis page Project Window Software Interface C 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 tg Analyze Markers x Q This analysis option is designed for Microsatellite and SNaPshot analyses Enter marker names separated by commas e o Cancel This option is only enabled when you are in the Project window Samples 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 GEnoWHeS SESE Sample Info Ctrl F 1 Raw Data Ctri F2 EPT Data Ctrl F3 BUIDEN Smia VAG Stale b Show Navigator Ctrl Shift N Project Window View Commands Item Description Enabling Samples Ctrl Shift 1 Switches to the Samples tab Always enabled Genotypes Ctrl Shift 2 Switches to the Genotypes tab Enabled when results are present Sample Info Ctrl F1 Switches to t
32. Files of type Java serialized file ser Cancel 6 7 Note Reference data and Size Standards used by exported Projects are not exported with the Projects Using the Project Window 3 25 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Export Project Window The elements called out in the figure in Export Project Window on page 3 25 are described in the table below Export Project Callouts Item Name Description 1 Toolbar These icons are used as follows Clicking moves display up one level in main pane Clicking moves display to Home level in main pane This level is usually Profiles lt user gt cx Clicking creates a new folder at the present directory level b b 0m0 bene Clicking presents a list of the contents of the selected folder Clicking presents details of the selected folder 2 Drop down directory menu Select drive letter and or folder 3 Folder file display pane The contents of the disk or directory are selected in the toolbar and the directory menu is displayed here Save button This button saves the exported project 5 File name field Enter a file name to use for saving your data Files of type field This field is a display filter for files folders are always shown Use this drop down menu to select the file extensions for display All Files
33. 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 x Ageia Het 3730 DS 33 Install BEIECh Sanne o Cancel Standard Parameters tep 1 2 3 Select the appropriate button Basic or Advanced or Classic The mode you select must be the same as the peak detection algorithm you use to analyze the samples in Analysis Method Using the Size Standard Settings 10 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To specify size standard parameters Step Action 4 If using the Classic mode a Select a dye color from the drop down list b Select an Analysis Method from the drop down list Select Dye and Analysis Method c Click the Select Sample button to open the Select Sample window and select the sample file that contains the dye standard you want to use as the template gy Select Sample 7 Fiwcsatene 2 E e eX E C006_E019_Run_C006_2002 05 10_205_07 fsa a CO06_E029_Run_C006_2002 05 10_205_05 fsa C006_E039_Run_C006_2002 05 10_205_03 fsa E C006_E109_Run_C006_2002 05 10_205_01 fsa Prosan paraca x 10 6 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To specify size standa
34. Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Feature Description Ability to create bin sets for Bin definitions which may vary different instruments between instrument types can be stored as separate bin sets allowing simultaneous analysis of data from different instrument types in the same project Ability to correct failed in lane size Individual samples that fail size standards standard matching can be corrected as soon as they are incurred without redefining the size standard definition About GeneMapper Software 1 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com How GeneMapper The following figure shows how data flows through the GeneMapper Software Works software system ABI PRISM i Size Calling Allele Calling instrument or Sequence Collector i Size Matching GeneMapper Database Raw data from Called Alleles Peak Detection Automatic Bin Assignment Algorithm ABAA Allele Peak Determination using Multiple Allele Peak Algorithms MAPDA Automatic Bin Builder ABB GR1867b 1 8 About GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Hardware and Software Requirements Hardware and The following table describes th
35. Guaranteed 888 88 SOURCE www artisantg com Tools Menu The Tools menu provides access to the GeneMapper Manager Panel Manager and Table Setting Editor applications Help GeneMapper Manager Ctrl M Panel Manager Ctrl J Table Setting Editor Ctrl T Show Offsets Options Item Description Enabling GeneMapper Opens the GeneMapper Always enabled Manager Ctrl M dialog box See GeneMapper Manager on page 2 11 Panel Manager Opens the Panel Manager Always enabled Ctrl J dialog box See Chapter 4 Using Panel Manager Table Setting Opens the Table Setting Only enabled when Editor Editor dialog box See samples are present Ctrl T Table Settings Editor on Only allows editing of a page 6 4 currently selected Table Profile 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 17 C 16 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 17 Add Samples Tab C 18 Sequence Collector Tab C 21 Analysis Tab C 22 U
36. Guaranteed 888 88 SOURCE www artisantg com Tools Menu The Tools menu provides access to the GeneMapper Manager and the Plot Settings Editor Tools Ctrl M Plot Settings Ctrl T Item Description GeneMapper Opens the GeneMapper Manager to allow creation or editing Manager of various project settings Used to create a new Plot Setting Ctrl M profile See Chapter 2 Using GeneMapper Software for more information Projects Analysis Methods Table Settings Plot Settings matrices size Standards Name Last Saved Owner Description j Bing 2002 06 19 11 1 am Factory Provided 2 Microsatellite Det 2002 06 19 11 1 gm Factory Provided 2 SNP Default 2002 06 19 11 1 gm Factory Provided Import Export Delete Note New profiles can only be created through the GeneMapper Manager Using Plot Windows Samples and Genotypes 7 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Plot Settings Ctrl T Opens the Plot Settings Editor to allow the currently selected Plot Setting profile to be edited See Chapter 8 Using the Plot Settings Editor for more information Plot Settings Editor Column lv Sample File Sample Name Panel Off scale 7 24 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentat
37. Limited Product Warranty 0 0 0 0 eee eee teenies Glossary Index Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About GeneMapper Software Chapter Overview Introduction This chapter describes the ABI PRISM GeneMapper Software Version 3 0 the relationship of GeneMapper software to the current ABI PRism genotyping software and the requirements for installing and starting GeneMapper software In This Chapter This chapter contains the following topics Topic See Page Overview of GeneMapper Software 1 2 Hardware and Software Requirements 1 9 Registering GeneMapper Software 1 11 Obtaining Technical Support 1 12 About GeneMapper Software 1 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Overview of GeneMapper Software About The GeneMapper software v3 0 is a new release of GeneMapper GeneMapper software This release of GeneMapper software has additional features Software and enhancements similar to the functionality of ABI PRISM GeneScan and Genotyper analysis software GeneMapper software provides automated genotyping for linkage analysis SNP validation population genetics and human identification users Supported The GeneMapper software v3 0 supports data ge
38. 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 Project Window Software Interface C 29 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Save Project and The Save Project and Save boxes enable you to save projects to the Save Dialog Boxes database The first dialog box is presented for the Save command if 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 Gp Save Project x G Project name a The Save dialog box opens when you select the Save As command Project name Baye Dont Save Cancel The elements of these dialog boxes are described in the table below Save Project Elements 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
39. Panel column select the panel that references this data Panels must be created in advance of Auto Binning and should contain all marker information known Using the Size Standard Settings 10 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To size reference data continued Step Action 4 In the Size Standard column select the size standard used with your data 5 Analyze your samples Samples are now ready to be used for Auto Binning in the Panel Manager Sizing SNaPshot Whether creating SNaPshot bins manually or by using the Auto Panel Reference Data feature with SNaPshot Primer Focus samples reference data needs to be sized To size reference data Step Action 1 Import SNaPshot or Primer Focus reference samples into the Project window 2 In the Analysis Method column select an Analysis Method with the following features Analysis Type set to SNaPshot Binset set to none Peak Detection Algorithm set for your data 3 Set the Panel column to none 4 In the Size Standard column select the size standard GS120LIZ or the appropriate size standard for your data 5 If using Primer Focus samples set the Sample Type column to Primer Focus 6 Analyze your samples Samples are now ready to be used for creating bins in the Panel Manager 10 20 Using the Size Standard Settings A
40. Process Quality Values for more information B 2 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com A flowchart of the data flow in GeneMapper software is shown below Standard signal processing is applied to the data prior to being delivered to the GeneMapper algorithms The algorithms discussed here are shown in the raised portion Input sample gt AB PRISM 310 377 systems gt Multicomponenting ABI Prism 3730 3700 3100 systems Vv Baselining Vv Peak detection Vv Binning Vv Size matching calling v Caller 1 Caller n Arbitrator Vv Bin assignment Vv Final quality value determination Vv Report results to user Peak Detection This algorithm uses Basic Advanced or Classic mode to detect peaks Algorithm and process data See Basic Mode Peak Detection Algorithm Settings on page B 8 Classic Mode Peak Detection Algorithm Settings on page B 9 and Advanced Mode Peak Detection Algorithm Settings on page B 14 Software Genotyping Algorithms B 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Size Matc
41. Process Quality Values Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com PQV Flags on the The PQV flags on the Genotypes page Genotypes Page Column Description AE Allele Edit check box The check box displays unchecked immediately after analysis The check box is checked when the marker allele calls have been edited by the user 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 ADO Allele Display Overflow check box The check box is checked when the number of alleles the marker calls exceeds the number to display previously set by the user The user specifies how many alleles to display in the Table Settings Editor parameters The default is two There are six columns for each allele to indicate name size height area mutation and comments SHP Sharp Peak A flag Pass Ii or Check A is automatically displayed when there is a sharp peak present in the marker signal Analysis does not stop if problems are detected with these properties but the user should examine results flagged with Check values 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 This flag is used for Linkage Mapping Microsatellite dinu
42. Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Analysis menu commands continued Item Description Enabling Analyze Analyses all samples eligible for Enabled when Ctrl R analysis The symbol is shown Samples tab is visible 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 visible Samples Analyze All Analyzes all samples whether Enabled when Ctrl Shift R or not the Status column Samples tab is visible reports that the 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 sample Samples tab is visible Analyze Markers Dialog Box on page C 14 Analysis Allows the user to change the Enabled when a Method Editor settings for analysis sample with an analysis method is selected Size Standard Editor Allows the user 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
43. Q 0 0 0 0 0 NNNNNNNNNN NNN NN NNN einiol oi oioi oioi oioi oioi nininini ai n eleleleleleleleleleleleselelalealas a Columns 1 marker name dye color min ASR ASR allele size range actually the marker size range max ASR 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 a zero O 8 comments if none write none Notice that there are no extra rows between markers and panels 4 24 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 4 Microsoft Excel Booki Mm x File Edit View Insert Format Tools Data Window Help la x Demak z A g 0 jA J23 pe A B Cc D EFIG H z 1 Version GM v3 0 2 Kit type MICROSATELLITE 3 Chernistry Kit Microsatellite Tutorial none 4 Panel Tutorial Panel 9 none 5 DBS264 Blue 108 130 2 0 none 6 D6S1574 Blue 146 172 2 O none 7 D65276 Blue 201 233 2 0 none 8 D55408 Blue 249 285 2 0 none 9 D65308 Blue 326 354 2 0 none 10 D65267 Green 104 136 2 0 none 11 D6S292 Green 154 176 2 0 none 12 D65434 Green 201 245 2 0 none 13 D55426 Green 274 298 2 0 none 14 D551981 Yellow 115 125 2 0 none 15 D65257 Yellow 167 195 2 0 none 16 D65446 Yellow 212 234 2 0 none 17 D55
44. 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 entry Data Points fields in order to specify only a limited range to be button 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 fields drop down menu in order to specify only a limited range analyzed for All Sizes each sample Partial Sizes B 14 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Smoothing and The Smoothing and Baselining parameter options help to optimize peak Baselining Options size and eliminate noise from the baseline Item Description Smoothing Used to help reduce the number of false peaks option buttons detected by the GeneMapper 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 adj
45. Values PQV 1 4 process quality values PQV A 1 editing A 2 overview A 2 PQV flags on the Genotypes page A 7 PQV flags on the Samples page A 6 PQV values for specific applications A 4 process A 2 rules for PQV columns A 5 using the PQV system A 3 Project discussion of supporting utilities 3 3 Project Analysis menu commands C 12 Project Edit menu commands C 11 Project Edit menu dialog boxes Find dialog C 46 Sort dialog C 47 Project File menu Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com commands C 9 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 30 Project Help menu commands C 26 Project View menu commands C 14 Project window importance of 3 2 interface conventions 3 5 navigating in 3 7 parts of the main window 3 6 reformatting 3 5 tasks performed in 3 3 toolbar 3 12 viewing contents 3 14 Projects page using the 3 23 Q Quality Flags tab options 5 17 PQV thresholds 5 16 R Raw Data tab 3 9 registration card completing 1 11 S sample files converting from Mac to NT E 2 Sample Information source of 2 8 Sample Sheet how GeneMapper uses 2 8 sample sheet 2 8 Samples and Genotypes views applying a menu selection 3 19 purposes of tables 3 11 Samples view appearance C 2 columns C 3 Artisan Technology Group Quality Instrumentation rules for
46. Will be guided through this process in more detail by reading the ABI PRISM GeneMapper ID Software v3 0 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 Setup the GeneMapper software the first time for each analysis gen ie application Import or create panels and bins in the Panel Manager Create an analysis method suitable for your application with appropriate bin sets No bin set is needed if you are performing a sizing only application Create the appropriate size standard s if your data uses size standards other than those provided with GeneMapper software Setup options so that the same setup will apply the next time you use GeneMapper software if desired Convert any Macintosh generated sample files to the fsa format See Appendix E for details Set up Project i i i indon ior analysis Setup 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 3 3 Perform analysis and examine the results Initiate analysis Examine analysis results using the Plot window s Export results Export re
47. a Macintosh computer to a computer running the Windows operating system a file extension is 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 Installing To install the sample file conversion programs on a Macintosh Conversion computer Programs 8 Step Action 1 Insert the GeneMapper software CD ROM into 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 Locate and double click the CONVFOLD folder Inside this folder are two files CONVPROG HQX and README TXT which contain the installation instructions 4 Copy the CONVPROG HQxX file to your local hard drive by clicking on the file dragging the file over to the local hard drive icon and dropping it in 5 Decompress the CONVPROG HQxX file by dragging and dropping it onto a program called Stuffit Expander Note You can download a free version of Stuffit Expander from http www stuffit com expander E 2 Sample File Conversion Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Converting Macintosh Sample Fil
48. a sample 9 5 assigning to sample 9 16 choosing a data point range 9 10 9 11 considerations 9 7 creating a GeneMapper 9 12 definition 9 3 eliminating primer peaks 9 11 evaluating 9 17 evaluation procedure 9 17 example 9 15 generating a new 9 12 process diagram 9 8 process of creating a new 9 8 Guaranteed 888 88 SOURCE www artisantg com raw data display 9 10 sample files 9 5 steps to create 9 9 using different computers for data collection and analysis 9 18 using with data collection 9 18 viewing the raw data 9 10 when to assign to a sample 9 5 when to create 9 6 matrix files about 9 3 to 9 7 bad matrix files causes 9 19 to 9 20 choosing a data point range 9 10 to 9 11 evaluating the matrix file 9 17 new matrix file generating 9 12 to 9 15 process of creating new file 9 8 to 9 9 Multiple Allele Peak Determination MAPDA 1 3 Multiple Allele Peak Determination Algorithms MAPDA 1 4 O Open Project dialog box C 28 Options dialog box C 17 P Panel and Bin text files 4 23 Panel data hierarchy 4 3 Panel definition format of text file 4 24 Panel definitions for SNaPshot 4 28 Panel Manager Add Bincommand 4 13 Add Reference Data command 4 17 Auto Bincommand 4 19 Auto Panel command 4 19 automatic bin assignment algorithm 4 40 automatic bin builder process 4 40 Bin view 4 37 4 38 4 39 Bins menu 4 12 Delete Bincommand 4 14 Delete Bin Set command 4 16 Delete SNP Marker command 4 13 Duplicate Panel co
49. exolab org Copyright 2000 Intalio Inc All rights reserved JNIRegistry 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 09 2002 4335526 Rev B Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Contents 1 About GeneMapper Software Chapter OVerview osc s0sde ceran 1b4 peo ante Redes be da E ad das 1 1 Overview of GeneMapper Software 0 0 0 c eee eee eee eee 1 2 Hardware and Software Requirements 00 000004 1 9 Registering GeneMapper Software 0 000020 ee eee eee 1 11 Obtaining Technical Support 2 0 0 0 00 cee eee ee eee 1 12 Using GeneMapper Software Chapter Overview ois cceenee bepce eee ed kedeas DE Rae EN EA 2 1 Steps Required to Use GeneMapper Software 0 0 2 2 Installing and Registering the GeneMapper Software 2 3 Logging on to and Logging out of the GeneMapper Software 2 5 Import
50. 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 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 In This Appendix This appendix contains the following topics Topic See Page GeneScan 120 Size Standard D 2 GeneScan 350 Size Standard D 3 GeneScan 400HD Size Standard D 4 GeneScan 500 and GeneScan 500 250 Size Standards D 6 GeneScan Size Standards D 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneScan 120 Size Standard About This Size The GeneScan 120 Size Standard is useful for sizing fragments Standard between 15 and 120 base pairs It is used when analyzing SNaPshot samples How It Is Prepared All aspects of the preparation of the GeneScan 120 LIZ size standard are proprietary Each fragment contains a single LIZ fluorophore GeneScan 120 The following table lists the GeneScan 120 LIZ denatured molecular Molecular Lengths fragme
51. in the GeneMapper 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 41 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 C 38 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Ctri 1 GM Database Ctrl 2 SEQUENCE Colector Cit Add Samples View Menu Commands Item Description Enabling Files Opens the Files tab Always enabled Ctrl 1 GM Opens the GeneMapper Always enabled Database Database tab Ctrl 2 Sequence Opens the Sequence Enabled and visible if Collector Collector tab Sequence Collector is Ctrl 3 configured and you are logged into Sequence Collector Configured through the Options menu Project Window Software Interface C 39 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Files Tab The Files tab e
52. least 512 MB RAM for the best results About GeneMapper Software 1 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Special The following is a list of special considerations for running the Considerations GeneMapper software v3 0 The ABI PRISM 3100 Genetic Analyzers require the 3100 Data Collection Software version 1 0 and a minimum of 512 MB RAM The Applied Biosystems 3730 3730xl Genetic Analyzers require the 3730 Data Collection Software version 1 0 When using the GeneMapper software v3 0 with the ABI PRISM 3700 DNA Analyzer the software must be installed on a separate computer GeneMapper software v3 0 requires a single processor computer GeneMapper software v3 0 runs on Windows NT and Windows 2000 platforms only Conversion utilities are included for analyzing data from Maclntosh based sample files GeneMapper software v3 0 is not compatible with Xeon chips on upgrades from GeneMapper software v 1 0 2 The version of Oracle database in the GeneMapper software v3 0 is an embedded license for use by five named users only IMPORTANT To accommodate more than five users additional GeneMapper software or Oracle database licenses must be purchased When installing GeneMapper software v3 0 the user must log into the local machine and have administrator privileges GeneMapper software v3 0 can exist with other Oracle clients but not
53. linear 10 18 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Performing a Sizing Only Application Introduction General Sizing Only Application Sizing only applications quickly analyze data without generating genotype calls To perform a general sizing only application Step Action 1 Import samples into the Project window 2 In the Analysis Method column select an analysis method with the following features The appropriate analysis type selected for your data A Binset of none The appropriate Peak Detection Algorithm set for your data 3 In the Size Standard column select the size standard used with your data 4 Analyze your samples Samples are analyzed for sizing only no genotype calls will be made When viewing samples in the Plot window the Sizing Table displays information for all peaks detected Sizing To take advantage of the Auto Bin feature for generating marker bins for Microsatellite microsatellite data reference data needs to be sized Reference Data To size reference data Step Action 1 Import microsatellite reference samples into the Project window 2 In the Analysis Method column select an Analysis Method with the following features Analysis Type set to Microsatellite Binset set to none Peak Detection Algorithm set for your data 3 In the
54. of the contents of the selected folder Clicking presents details of the selected folder 2 Drop down directory menu Select drive letter and or folder 3 Folder file display pane The contents of the disk or directory are selected in the toolbar and the directory menu is displayed here Import button This button imports the selected project 5 File name field Enters name of selected project being imported Files of type field This is a display filter for files folders are always shown Selects the file extensions for display All Files shows all files Java Serialized file ser shows only files with the extension ser which is how GeneMapper database projects are exported Note There is only one preset file extension ser that can be used 7 Cancel button Closes the Import Project window without importing a project 3 28 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Panel Manager Chapter Overview Introduction This chapter describes the Panel Manager feature in the ABI Prism GeneMapper Software Version 3 0 and provides instructions on how to use it for microsatellite data Some information is provided regarding SNaPshot specific features however additional information can be found in the tutorial guide SNP Genotyping with ABI PRism GeneMapper
55. other Oracle servers The only exception to this rule is when the ABI Prism 3100 Data Collection Software version 1 0 and or the Applied Biosystems 3730 Data Collection Software version 1 0 is installed on the system 1 10 About GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Registering GeneMapper Software How to Register To register fill out the registration card included in this package and return it to Applied Biosystems and read the information in Chapter 2 Installing and Registering the GeneMapper Software on page 2 3 For Applied Biosystems technical support telephone and address information see Obtaining Technical Support on page 1 12 Note Installation privileges are available only if you have returned your registration card About GeneMapper Software 1 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Obtaining Technical Support Applied A services and support page is available on the Applied Biosystems Biosystems Web site To access this go to Website ittp www appliedbiosystems com and click the link for services and support At the services and support page you can Search through frequently asked questions FAQs Submit a question directly to Technical Support Order Applied Biosystems user documents MSDSs certificates of analysis and other related
56. peaks with those peaks whose amplitude above 50 are heights are at least the analyzed and display in Peak Amplitude the tabular data Threshold for that dye Lower amplitude peaks Set the dye amplitude still display in the threshold at a level that 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 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 Software Genotyping Algorithms B 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description For example Polynomial Degree Sets the degree of the polynomial Min Max setting 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 w
57. process Ease of use One button operation is provided with fully integrated multi step size calling allele calling and intelligent data management using the GeneMapper database Unique features of the GeneMapper software v3 0 Unique Feature Description Multiple Allele Peak Determination For non forensic data the MAPDA are used Aaprhas MARDA For forensic data the Human Identity Caller HIC algorithm is specifically optimized to handle data containing tetranucleotide repeats and allelic ladders Automatic Bin Builder ABB After allele peaks are called using MAPDA the ABB is used to Automatic Bin Assignment create the bins optimizing the distances between bins and the Algorithms ABAA precise locations of bin centers The ABAA then completes the allele calling process by assigning allele peaks to their corresponding bins Process Component Based After bin building and allele assignment bin quality values are Quality Values PQV assigned to these bins based on the success rate of the bins to locate allel k i ion See Appendix A for more ocate allele peaks correctly from the sample file collection information The PQV system facilitates fully automated operation with only questionable allele calls those with minor component related problems requiring any intervention by the user Bad samples defined as being below the threshold of acceptability can be discarded without any examination
58. project specific folders Clicking a kit folder displays the Panel table J Panel Manager 8 5 LMS HD5 V2 5 9 Panel01 HD5 2 5 C Panel02 HD5 2 5 The Panel folder icon represents a set of markers Clicking a panel folder displays the Marker table For SNP kits clicking a panel folder also displays a plot view 3 6 Panel Manager E LMS HD5 2 5 Panel01 HD5 V2 5 D1S2797 D18249 The Marker icon represents an individual marker Clicking a microsatellite marker displays the Bin view Clicking an SNP marker displays the Bin table Using Panel Manager 4 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Manager Window Commands Introduction The tables in this section describe the following Panel Manager views and menus Topic See Page File Menu 4 6 Edit Menu 4 11 Bins Menu 4 12 View Menu 4 20 File Menu The File menu offers options to the user to create or export kits duplicate import and export panels and import and export bin sets AGE Edit Bins View New Panel Ctrl N DUB Cate Farrel libot Panels ithe sy Import Bin Set Ctrl Shift B Export Panels Ctrl E Export Bin Set Ctrl Shift E Export All Kits Ctri K Ctrl P Print New Kit Panel Marker The New Kit Panel Marker command changes its name and function depending on the item selected in the Panel Manage
59. the active tab tot GeneMapper Project Microsatellite Project File Edit Analysis View Tools Help Aea BA Mil 3 Project SNaPshot 4 Linkage 02_1004 03_1005 04_1003 05_1004 06_1005 07_1003 4 EG Info Raw Data EPT Data Table Setting applied Biosystems Appli z m og Sample Information Sample File Sample Origin Path Status File Source 01_100301 fsa Need to be analyzed Local drive Error Message None Message D GeneMapper Examples SampleFiles Linkage 01_100301 f3sa Progress Status 3 8 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The Info tab displays sample file information such as sample information run information data collection settings capillary information gel information error messages 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 gp GeneMapper Project Microsatellite Project oO x File Edit Analysis View Tools Help ech BS Bl MuE Project Info Raw Data EPT Data C1sNaPshot a4 100301 fsa B f Linkage 4400 1838 2276 2
60. to exit and reenter the database host information Select the appropriate machine type from the drop down list 5 Click OK 2 6 Using GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneMapper The following rules apply to access the GeneMapper software Software Access 4 The first time the GeneMapper software application is launched Rules after installation user registration is required Once registered all users can read write and edit everything in the database except predefined content because there are no access privileges Sequence Collector access information is specified in the Options dialog box in the Tools menu Only one user can access a database at a time Database connections can only be made across computers using GeneMapper software v3 0 and not earlier versions of the software Logging Out Close the GeneMapper software in one of three ways Click File gt Logout Click File gt Exit or Click the Close button x If you close GeneMapper software with a new or blank Project window or no changes in your Project the Save Project logout dialog box does not open When you log out of the GeneMapper software you have the option of saving or discarding the changes you made since the last time you updated the project oy Save Project Ea N Do you want to save changes i no cance If y
61. weighted on a scale of 0 to 1 with 0 being of no importance and 1 being of the highest importance Most defaults are set at 0 5 except for Out of Bin Allele which is set at 0 8 These defaults emphasize the point that if a peak falls outside a bin it should have a lower genotype call The Default button will reset these calls to the default numbers discussed in the Peak Quality Tab section on page 5 14 Analysis Method Editor Microsatellite General Allele Peak Detector Peak Quality Quality Flags Quality weights are between 0 and 1 Quality Flag Settings Spectral Pull up fos Control Concordance fos Broad Peak fos Low Peak Height fos Single Peak Artifact jos Off scale fos 5 Sharp Peak bs PeakHeightRatio 0 5 One Basepair Allele 0 5 _ OutofBin Allele 08 Split Peak ps PQY Thresholds Sizing Quality From fo 7s to 1 0 From 0 0 to 0 25 Genotype Quality From 7s to 1 0 From 0 0 to fo 26 Factory Defaults oc cna PQV Thresholds 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 See the figure on page 5 16 Anything that is not labeled Pass or Low Quality will be labeled as check 5 16 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Options The Quality me
62. when you want to ignore a peak for the standard definition Press the tab key to automatically move to the next size standard peak Click OK to save and close the Size Standard Editor or click Cancel to close without saving Using the Size Standard Settings 10 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 the size calling curve To open the Size Match Editor window click Analysis on the toolbar in the GeneMapper Project window and select Size Match Editor Edit View Tools x Eg Size Matches size Calling Cure Sizing Quality 1 0 Override SQ aua 550 1100 1650 2200 2750 3300 3850 4400 E E When the Size Match Quality indicator shows a marginal or fail state you can use the Size Match Editor to help adjust the in lane size standard to make it valid Typical problems include the following conditions Peak shift The algorithm does not correctly identify a peak resulting in invalid positions for other size standard peaks Missing peak The algorithm does not identify a peak Extra peak The algorithm identifies a peak that is not a size standard peak but a spike or pull up peak 10 12 Using the Size Standard Settings Artisa
63. 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 4 Apply additional project settings prior to analyzing the sample files 9 16 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Evaluating the Matrix Introduction After creating a new matrix and assigning it to select sample files evaluate the quality of the matrix The quality of the matrix has a direct impact on the quality of the results data Procedure To evaluate the matrix Step Action 1 Analyze the sample files used to make the matrix 2 Display results data for all the Dye Matrix Standard sample files in the plot window showing only electropherogram data For each displayed sample file you should see the following You should see If not that the only visible peaks represent you probably have a bad matrix the color of the Dye Matrix Standard run in that lane or for that injection p l and correct problems with bad pel eae matrices see Causes for Bad all other lines should be relatively Matrices on page 9 19 flat indicating that the matrix properly compensated for the spectral overlap For instructions on how to identify For example for the blue matrix standard sample fi
64. 0 Calculation 2 Generating a New Matrix 9 12 3 Assigning the Matrix to Samples 9 16 4 Evaluating the Matrix 9 17 5 Using the Matrix with Data Collection 9 18 6 Causes for Bad Matrices 9 19 Creating and Evaluating a Matrix 9 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Choosing a Data Point Range for the Matrix Calculation Introduction 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 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 Viewing the Raw To view raw data Data Step Action 1 Open a new GeneMapper Project 2 Import Samples by selecting File gt Add Samples to Project 3 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 In the navigation pane expand the folder containing sample files Select a sample file
65. 0 402 144 128 138 150 4162 474 196 198 210 222 234 24 258 DS33 LMS fsa Dss3 INSTALL ps 33 m 4 66 78 90 4102 144 126 438 450 4162 174 186 198 210 222 234 24 258 5 1992 1511 na Headers Turned Off x File Edit View Tools Alleles Help posan EQ Pse M AA WE Tcl masis 90 402 1444 142 438 450 462 174 1896 4198 210 222 234 24 258 90 402 144 1428 438 450 462 474 196 4138 210 222 234 24 258 Marker Range Ctrl R Controls whether or not the colored marker bar is displayed at the top of an electropherogram 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 Using Plot Windows Samples and Genotypes 7 21 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Marker Margin Opens the Marker Margin dialog box to allow the user 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 gy Marker Margin x Enter marker margin in basepairs FFF Cancel 7 22 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation
66. 0 has user defined columns 1 3 in the sample sheet Using Sample The sample sheet information is essential for associating the nature of Sheet Information sample fragments with individual dye lanes and tables in the GeneMapper software For example the following figures show how the GeneMapper software incorporates information entered in the Sample Name Color Info and the Color Comment fields of a sample sheet into GeneMapper 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 software with the necessary information 2 8 Using GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sample Sheet GeneScan1_1D test for GM GeneMapper Software Project Table Microsatellite Default aE Projeci k ETLinkage Status Sample File Sample Name Comments Sample Type Analysis Method Size Standard Matrix Ri 2 M 02_100405 fsa 100405 None Sample None None Porkchop ROX Matrix Li 3 Be 03_100501 fsa 100501 None Sample None None Porkchop ROX Matrix Li 4 be 04_100302 fsa 100302 None Sample None None Porkchop ROX Matrix Li User defined Read from Sample File Find Command You can increase the utility of the Find Ctrl F command in GeneMapper software by carefully planning the format of the Using GeneMapper Softw
67. 1 0 Override SQ 3407 3597 3787 3977 3 16 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Access GeneMapper Application Windows continued To see the Select And GeneMapper software displays the Table Setting Tools gt Table Table Setting Editor Editor Setting Editor x or press General Samples Genotypes Ctrl T pme Table Settings 1 ie Show Column Filtering Content o If samples 1 E Jerus Show All Records NA have been 2 m Sample File Show All Records imported you 3 Sample Name Show All Records can click the 40 Sample ID Show All Records 5 Comments Show All Records icon 6 Mv Sample Type Show All Records N A 7 M Analysis Method Show All Records 8 m Panel Show All Records Note To 9 Size Standard Show All Records create a new 1 Matrix table select Tools gt GeneMapper eane Manager Then select the Table Settings tab and click New Plot window a sample or Plot Window results row and then either select lo Analysis gt crosan Ei Panes gt ee el Display Plots press Ctrl L or click the icon i A Wee Male Sample File Sample Name Pand os sQ DS33 INSTALL DS 33 427 159 Using the Project Window 3 17 Artisan Tec
68. 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To create a GeneMapper software matrix continued Step Action 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 Matrix Settings SSS 7 Select the Matrix Standard Sample File Number of Dyes _B_ No File Selected for B Data Startat 1000 G No File Selected for G Data Start At fiooo Y No File Selected for Y Data Start At foo R No File Selected for R Data Start At foo O _ No File Selected for o Data Startat jooo Points fi 00000 Create 5 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 6 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 9 11 7 Enter the total number of data points to include to calculate the matrix in the Points field In most cases leave the default value unless you must exclude a portion of your data because of artifacts or bleed through Note You must have at least five peaks to make
69. 41 Goe t 5354 1045 Electropherogram showing four resolved peaks detected as two peaks Software Genotyping Algorithms B 25 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Effects of The figure below shows the data presented in the figure above see Reducing the page B 25 re analyzed with a window size value of 10 and polynomial Window Size Value degree value of 5 and Increasing the Polynomial Degree Value w A J820 1890 1840 1850 1800 1870 1980 1890 1900 1910 1020 1990 1940 1950 1900 1970 1980 1990 2000 2010 2020 2030 2040 3R 17 611 335 60 188 1126 1916 3R 18 a8 337 54 859 4ag0 1525 3R 19 516 339 50 1402 10486 1935 3R 20 E E E a a A TE 3R 21 521 342 82 63 203 1953 Electropherogram showing all four peaks detected after reducing the window size value and increasing the polynomial degree value B 26 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Optimizing Peak Detection Sensitivity Example 3 Effects of Extreme The figure below shows the result of an analysis using a peak window Settings 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 ey 4010 4020 4030 4040 4060 4060 4070 4080 4090 4100 410 4120 4130
70. 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com PQV Flags on the The PQV flags on the Samples page Samples Page PQV Column Description SFNF Sample File Not Found A flag Pass Ii or Check A is automatically displayed when no sample file is found Not editable MNF Matrix Not Found A flag Pass Ii or Check A is automatically displayed when no matrix file is found attached to the sample file Not editable SNF Size Standard Not Found A flag Pass Ii or Check A is automatically displayed when no size standard is found Not editable OS Offscale A flag Pass Ii or Check A is automatically displayed when there is offscale data in the signal Not editable Indicates an offscale result in the size standard region of the signal SQ Sizing Quality A value Pass i Check A or Low Quality is automatically displayed to indicate the sizing process quality 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 Not editable 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 A 6
71. 641 Yellow 299 339 2 0 none 18 D55433 Red 69 99 2 0 none 19 D55422 Red 118 140 2 0 none 20 D55406 Red 168 196 2 0 none 21 D55400 Red 221 243 2 0 none 22 D65309 Red 307 333 2 0 none 1414 Mh Sheet1_ Sheet2 sheets 7 tal Res CI nn S r 1 2 3 4567 8 Columns 1 marker name 2 dye color 3 min ASR ASR allele size range actually the marker size range 4 max ASR 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 a zero O 8 comments if none write none Notice that there are no extra rows between markers and panels Using Panel Manager 4 25 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Bins Definition The examples below show bins defined in the format used by Example GeneMapper software v3 0 in a tab delimited text file txt or an Excel spreadsheet saved as a txt file Note The 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 3730 DS 33 Install Bins txt WordPad o Oj xj File Edit View Insert Format Help pela SI al lel Version GH v 3 0 Chemistry Kit GeneScan Installation Standards BinsSet Name 3730 D5 33 Install Bins Panel Name
72. 6506E 02 T Al 1 608974E 01 A2 2 041442E 05 A3 1 887158E 09 R 2 1 000 250 200 150 100 Size Calling Curve 3rd Order Least Squares T T T T 2500 3000 3500 4000 Data Point 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 software v3 0 Software Genotyping Algorithms B 33 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Cubic Spline By definition the Cubic Spline method forces the sizing curve through Interpolation all the known points of the selected size standard Although this Method nforement produces exact results for the values of the standards themselves it does not compensate for standard fragments that may run anomalously ELI 11eP2 11 Best Fit 2nd Order Curve AO 9 771623E 01 Al 1 220838E 01 A2 4 856885E 06 R 2 1 000 Size Calling Curve Cubic Spline Interpolation T T T T T T T T 1000 1500 2000 2500 3000 3500 4000 4500 Data Point TAAS A T Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper software v3 0 Possible Local Sizing Inaccuracy Mobility of any DNA fragment can be affected by its sequence and by secondary and tertiar
73. 714 3152 3590 4028 4466 4904 5342 5780 6218 6656 7094 E Table Setting fApplied Biosystems Appli 02_1004 03_1008 04_1003 05_1004 06_1005 07_1003 E 2 scan 4583 rfu 2760 Progress Status AA There are several plot scaling features in the Raw Data view 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 Return to a full scale plot by double clicking in the label area again The plot window is restored to full scale You can scale the Y axis to maximum Y or a user defined value by selecting Y Axis Scale from the View menu Using the Project Window 3 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Use the Raw Data view to evaluate the following problems or noise in the baseline that could result in poor size calling 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 Qp GeneMapper Project Microsatellite Project _ ol x File Edit Analysis View Tools Help eos a BS i Lil i W W Tabie Setting Applied Biosystems Appli z fin 8
74. Always enabled Export button xml file Displays a dialog box for Exporting selected objects Enabled when one or more objects are selected Delete button Deletes the selected object s Done button Closes the GeneMapper Manager Using Table Settings 6 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Table Settings Editor Introduction 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 The Table Settings Editor window contains the following three tabs General Samples Genotypes The General Tab The General tab provides the capability to give a name and description to a new profile Note You cannot change the name of an existing profile only the description amp Table Setting Editor x Samples Genotypes ew Profile 6 4 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The Samples Tab The Samples tab provides the capability to filter columns in the Samples table of the project window g Table Setting Editor Filtering ses otowanrecoas NA m Sample File ShowAN Records m Sample Name ShowAN Records F_ Sammie fenon _ oommens C _ Siete _ orowanrews WA _ Awsswebed l
75. 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 Bin 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 4 38 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 location 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
76. DS5 33 Marker Name D205119 113 112 92 0 4 119 118 75 0 4 Marker Name D951690 239 239 3 0 4 0 241 241 3 0 5 0 Marker Name D55644 65 84 94 0 4 oO 97 96 95 0 4 Qi Marker Name D55424 218 218 24 220 220 18 0 Marker Name D95288 137 137 45 0 146 146 2 0 4 Qo Marker Name D185462 303 303 05 D ARUN 1 2 3 4 Columns Rows 1 bin name 1 GM version number 2 bin center 2 Kit name 3 bin left offset 3 bin set name 4 bin right offset 4 panel name 5 marker name 4 26 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com fa Microsoft Excel Book1 loj x E fie Edit wiew Insert Format Tools Data window Help x Demla A 2 i or 2IA A B Ch Tr Lee GM v3 0 1 2 Chemistry Kit GeneScan Installation Standards j 2 3 BinSet Name 3730 DS 33 Install Bins i 3 4 Panel Name DS 33 i 4 5 Marker Name D205119 5 6 113 112 92 0 4 0 5 119 118 75 0 4 0 5 8 Marker Name D9S1690 9 239 239 3 0 405 10 241 241 3 0 5 05 11 Marker Name D55644 E 85 84 94 0 4 0 4 13 g7 96 95 0 4 0 5 14 Marker Name D55424 15 218 218 24 0 4 0 5 16 220 220 18 0 5 0 5 47 Marker Name D9S288 18 137 137 45 0 4 0 4 19 146 146 2 0 4 0 4 20 Marker Name D18S462 21 303 303 05 0 4 04 I4 4 gt gt i Sheet1 Sheet2 Sheets f gt see el ee 1 2 3 4 Columns Rows 1
77. Determination Quality Values PQVs PQVs are reported by data analysis and are an Algorithm 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 PQV x PQV x xPQVj PQVs are weighted from 0 to 1 The actual value of a PQV in the equation is 1 minus the weight 0 weight no effect on the final GQ calculation 1 minus 0 1 therefore no change to GQ value 4 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 failed Between 0 and 1 the higher the value the greater the impact on GQ PQV filtering is controlled by the threshold set by GeneMapper 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 Software Genotyping Algorithms B 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Basic Mode Peak Detection Algorithm Settings Overview The Basic peak detection algorithm is used as the default algorithm by GeneMapper software v3 0 Basic Mode uses t
78. 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 Edit View Files GM Database Seguente meea Samples To Add El Linkage i a 01_100301 fsa i a 02_100405 fsa i a 03_100501 fsa a 04_100302 tsa j i a 05_100406 fsa i a 06_100502 fsa fa 07_100305 tsa 29 SNaPshot xl Add To List gt gt i __ dptions i C 36 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To add samples to the Project Step Action 1 Click Bj Add Samples To Project on the Project window toolbar Add Samples to Project window opens to the last tab viewed by the user default is Files tab Sample files can reside on the local hard drive s mapped network drive s and removable media drives If Sequence Collector is configured and you are logged into Sequence Collector the Sequence Collector tab is enabled 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 Click the Add
79. Instrumentation Guaranteed 888 88 SOURCE www artisantg com Users Tab Elements continued Element Description New User button Opens the New User dialog box ty New User x New user name Password Confirm Password To add a user name for GeneMapper software click New User to open a dialog box and then 1 Type a user name into the dialog box 2 Enter a password using only alphabetic characters 3 Confirm the password 4 Click OK to assign the user name and close the dialog When you select this user name on launching GeneMapper software the name is used to identify the preferences set by the user 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 software installation and is visible to all other users The names are converted to lower case and passwords are not case sensitive Project Window Software Interface C 25 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Users Tab Elements continued Element Description Change Opens the Change Password dialog box and allows you Password to change the password for the selected user Change password for user gm Password Confirm Password Help Menu The Help menu provides access to online help and to the About GeneMapper software window
80. M 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 Instrument Type From sample sheet not editable Instrument ID From sample sheet not editable Run Date and Time From sample sheet not editable REF Reference data 3 indicates that the selected sample is defined as reference data in the Panel Manager SQI Sizing quality invalidated checkmark indicates that sizing quality value is 1 0 User Defined 1 3 Editable free text Also from the vertical bar enabled in the sample sheet fields See Chapter 2 Using GeneMapper Software Note Refer to Appendix A for PQV information C 4 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 continuous 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 Cirl click the Row Header box Project Window Software Interface C 5 Artisan Technolog
81. OYysdeNS BuidAyouey dNS dNS pue uoneajyuap UEWNH AIH uaeso1N Budde seyun WI siskjeue yore JO pasn ase sanyen yorym smous giqe Bulmo jo suoneyddy Iyp ds y seshjeue o ds 104 pasn aie UOIJEWOJU BJOW JO y BHed s s njer AOd euL 107 SNBA AOd A 4 Process Quality Values Guaranteed 888 88 SOURCE www artisantg com Artisan Technology Group Quality Instrumentation Rules for PQV In gaining familiarity with the PQV columns on the Samples page and Columns Genotypes page keep the following rules in mind 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 the user should examine results flagged with Check values Holding the cursor over a column header displays a ToolTip identifying the full name of the column the default names are often acronyms PQV results are reported in the column labeled SQ on the Samples page and GQ on the Genotypes page 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 Process Quality Values A
82. Plot Settings Editor 8 4 Using the Plot Settings Editor 8 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Plot Settings Tab Introduction The Plot Settings tab enables you to create and edit plot setting profiles for viewing your data in the Samples and Genotypes plot views Default Profiles GeneMapper software v3 0 contains the following standard Applied Biosystems profiles Microsatellite Default SNP Default Sizing view is similar to the view in the ABI PRism GeneScan software Plot Settings Tab The Plot Settings tab contains a list of Plot Setting profiles in the database and is used to manage the contents of the Samples and Genotypes Plot views by providing the following menus and interface elements To display the Plot Settings tab select Tools gt GeneMapper Manager gt Plot Settings meee x Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Name LastSaved Owner Description a Sizing 2002 06 10 17 0 gm Factory Provided 2 Microsatellite Default 2002 06 10 17 0t gm Factory Provided 3 SNP Default 2002 06 10 17 06 gm Factory Provided 8 2 Using the Plot Settings Editor Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The Plot Settings tab contains the following items Item Description Name column Plot setting name Las
83. Pshot Microsatellite OK Cancel 5 6 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com General Tab In the General tab enter a name and description for the analysis method Analysis Method Editor Microsatellite Note The Analysis Type reflects the choice selected in the New Analysis Method dialog box Allele Tab In the Allele tab set a number of parameters controlling allele calling including Bin Set Marker Repeat Type for Microsatellites SNP Cut Off value for SNaPshot Using Analysis Methods 5 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Analysis Method Editor Microsatellite Bin Set Bins are a way of matching your data to the allele definitions contained in the panel assigned to the data Bins are the locations in which you expect an allele to display Alleles are defined in Markers as a size range bin centered around the average size in base pairs for example 101 5 0 4 bp If your sample has a peak that falls within a bin the allele in your sample file is called with the identifier assigned to that allele bin A bin set must be chosen 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 ma
84. R_Panels_v3 Microsatellite none 4 LMSHD5 V2 5 4 LMS HD5 Microsatellite none EJLMS LD20 5 LMSHDS V2 5 Microsatellite none C LMSMD10 V2 5 6 LMS LD20 Microsatellite none 7 LMSMD10 V2 5 Microsatellite none amp 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 Using Panel Manager 4 33 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Table The table below lists the columns in the Panel table Columns Note All columns are resizable Column Name Description Panel Editable Contains the name of the panel Cell accepts Name alphanumeric characters restricted for any invalid symbol characters includes all Microsoft Windows invalid characters Note When 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 Editable Contains comments regarding the panel Cell accepts alphanumeric characters Name Rule for Panel names must be unique within a kit and across kits Panels 4 34 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Marker Table View Introduction Selecting a panel in the navigation pane displays the Mark
85. 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 button Adds the selected items from the Results field to the Samples to Add field Samples To Add field Contains the folders and files that the system will add 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 Project Window Software Interface C 43 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sequence Collector The Sequence Collector tab enables you to choose sample files that Tab are stored in a Sequence Collector The tab uses a search interface to extract samples that match your criteria The Sequence Collector tab opens only when a connection to Sequence Collector exists gt Add Samples to Project i Add ee Anay The following table describes elements of the Sequence Collector tab Sequence Collector Tab Elements Element Description Search Criteria This table contains the search criteria from which to extract samples from the Sequence Colle
86. Show All Records Show Pass Show Check Show Low Quality 6 14 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Plot Windows Samples and Genotypes Chapter Overview Introduction This chapter describes how to examine and interpret electropherograms within the ABI Prism GeneMapper Software Version 3 0 Samples and Genotypes plot windows and explains the various electropherogram settings and interactions in the plot window menu items In This Chapter This chapter contains the following topics Topic See Page About the Plot Window 7 2 Plot Window Toolbars 7 4 Plot Window Menus 7 5 Using Plot Windows Samples and Genotypes 7 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About the Plot Window Introduction The Samples and Genotypes plot windows allow you to visually assess your data If genotype calls were made you can also view and edit allele calls assigned by the GeneMapper software algorithms Purpose of the Plot Samples Plot Window Window The Samples plot window allows you to View electropherograms on a per sample basis View a sizing or genotypes table for sample s shown Overlay all samples to determine size standard quality Edit markers and bins View the relationship between controls and samples gt gt gt gt
87. Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sort Dialog Box The Sort dialog box shown below allows sorting of sample files results etc by ascending or descending order in up to three steps ia Sort x Sort by Ascending Sample File bull E z i Descending Then by Ascending Sample File Bll a i 2 Descending Then by Ascending Sample File Descending The Sort dialog box has three entry fields for entering sort criteria the Then by fields allow successive sorting by two other criteria Project Window Sort Elements Element Description Sort by Sort box containing the column which will be sorted Then by Ascending Option buttons for choosing if the 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 Project Window Software Interface C 47 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneScan Size Standards Appendix Overview Introduction The ABI Prism GeneMapper Software Version 3 0 comes with several ready to use GeneScan size standard definition
88. The following information is provided in this section for the Samples view Topic See Page Appearance of Samples View C 2 Samples View Columns C 3 Samples View Features C 5 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 gp GeneMapper Project Microsatellite Project ol File Edit Analysis View Tools Help ee SG BB ih ll HE gt Tablesetting ERgeeem Project Samples Gennijnes Linkage T fat EI SNaPshot Status Sample File Sample Name Comments Sample Type Analysis Method Panel Size 9 8 Pe 28 3 fsa 2 None Sample SNaPshot Default None clasg 5_E03_09 fsa 5 None Sample Sample Sh shot Default None SNaPshot Default 6_FO3_11fsa 6 iiz _ 04_06 fsa liz Sample Sample SNaPshot Default SNaPshot Default analysis Completed ed Note You can hide and show columns in the table using Table Setting Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Samples View Columns The columns of the Samples view are explained in the table below Note The following image is shown with the navigation pane closed amp
89. _Control COfiler_v3 D78820 Y 10 11 272 24 276 28 678 0 725 0 5612 0 6331 0 GeneMz Genemz i E 8 CO_Ladder fsa CO_Ladder COfiler_ v3 D381358 8 12 13 111 35 115 49 1611 0 1589 0 10777 0 10110 0 GeneMe GeneMz E X E 9 CO_Ladder fsa CO_Ladder COfiler v3 D16S639 B 5 8 229 45 241 44 1575 0 1658 0 11731 0 12612 0 GeneMe Genems E X E 4 Laddar inn Lada filar u2 een I hoo es angas Lanonn hasana hacazrenhannnan aa Canan m L I m Analysis Completed C 6 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotype View Columns The table in the Genotypes view can display some of the columns in the Samples view see table on page C 3 plus these unique columns Genotypes View Columns Column Description Marker Not editable name of the marker associated with the record created in the Panel Manager 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 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 f
90. a marker bin as follows For microsatellite data Step Action 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 Edit the bin information in the dialog box that opens For SNaPshot data Step Action 1 Select a SNaPshot 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 on the bin and select the Edit Bin option 4 Edit the bin information in the Edit SNP Marker dialog box that opens 4 14 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Name 313 Location 2 79 Left offset 0 40 i Right offset io 40 I Mutant Bin Cancel New offset default New offset default 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 fr
91. a matrix 8 Click Create to generates a new matrix 9 Click OK to save and close the Matrix Editor dialog box 9 14 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Matrix Example The following is an example of the matrix results showing the values used to calculate the overlap correction Matrix Result B G Y R 0 B h 0000 0 2009 0 1176 0 0319 0 0385 G 0 9422 1 0000 0 8531 fo4g77 bars if a s628 0 6374 1 0000 h 9224 28925 R o 2607 0 3527 0 7938 hooo l AT 0 oo4o4 0 0962 0 1992 loan l 0000 Ot 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 Creating and Evaluating a Matrix 9 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Evaluating the Matrix on page 9 17 Procedure To assign a matrix to samples Step Action 1 Open or create a GeneMapper Project containing the samples
92. able columns will display correctly in text format 3 22 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Projects Page Using the Projects Page The GeneMapper Manager Projects page shown below is a utility which is used to rename save as import export or delete projects To display the Projects page select the GeneMapper Manager from the Tools menu and select the Projects tab G GeneMapper Manager oje 45i 8 1 oject 2 Casework Project 06050 2002 06 05 15 2 BO 3 HID Casework 062602 2002 08 01 15 3 om BO 4 Database Project 2002 07 01 09 1 6M fh Select the project you would like to change and click the appropriate button If you click the Delete button the Project Deletion dialog box displays Click No if you do not want to delete the project Click Yes only if you want to delete the project permanently E GeneMapper Deletion Alert Q Using the Project Window 3 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Projects Tab The Projects Tab allows you to view information about the projects stored within the GeneMapper database and perform certain functions such as renaming and exporting of projects Note The only method of backing up your projects is to export them Description of items on the Projects tab Item Description
93. above contains the following items used for printing Item 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 Allis the only option available when working with tables Number of Copies Select the number of copies to print Collate options Check box enables collating of copies Project Window Software Interface C 33 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 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 Edi
94. ality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Table Settings Chapter Overview Introduction This chapter provides information about the Table Settings tab in the GeneMapper Manager window of the ABI Prism GeneMapper Software Version 3 0 and describes how to use the features of the Table Settings Editor In This Chapter This chapter contains the following topics Topic See Page Table Settings Tab 6 2 Table Settings Editor 6 4 Using Table Settings 6 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Table Settings Tab Introduction Use the Table Settings tab to perform the following tasks Create new profiles Hide and show table columns Filter the table entries rows in Project windows Default Table GeneMapper software v3 0 contains the following standard Applied Settings Biosystems profiles Default SNP Default Microsatellite Default Default is a general profile SNP Default is intended for SNaPshot samples and Microsatellite Default is intended for Microsatellite samples These profiles can be used as models for customizing new profiles Table Settings Tab To display the Table Settings tab select Tools gt GeneMapper Manager gt Table Settings LI x Projects A
95. alphanumeric characters from the keyboard The field is validated if the marker name is unique in the panel Note Marker names are case insensitive i e D1S83 is the same as d1s83 Dye Color Editable Contains the color names blue green yellow red orange The field is validated if it is one of these text strings which are not case sensitive Min Size Editable Contains the starting base pair number for the allele size range Cell accepts numeric characters from the keyboard The range defined by the Minimum Size parameter should include the stutter peaks for the smallest allele Max Size Editable Contains the ending base pair number for the allele size range The cell accepts numeric characters from the keyboard Control Editable Contains a comma separated list of control allele Alleles names in the marker No validation Marker Editable Indicates a di tri or tetra nucleotide repeat R t open 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 Marker Editable Contains the ratio of stutter peak to main peak Specific Sets the maximum value of stutter for a specific marker as Stutter Ratio opposed to using a global value for all markers Comments Editable Contains comments regarding the marker Cell accepts alphanumeric characters from the keyboard Ladder Editable Comma separ
96. and Advanced algorithm settings In this Appendix This appendix contains the following topics Topic See Page GeneMapper Software Genotyping Algorithms B 2 Basic Mode Peak Detection Algorithm Settings B 8 Classic Mode Peak Detection Algorithm Settings B 9 Advanced Mode Peak Detection Algorithm Settings B 14 Peak Detection Polynomial Degree and Peak Window Size B 19 Parameters Optimizing Peak Detection Sensitivity Example 1 B 23 Optimizing Peak Detection Sensitivity Example 2 B 25 Optimizing Peak Detection Sensitivity Example 3 B 27 Peak Detection Slope Threshold for Peak Start and Slope B 28 Threshold for Peak End Parameters Slope Threshold Examples B 30 Size Calling Methods for Classic and Advanced Modes B 32 Software Genotyping Algorithms B 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneMapper Software Genotyping Algorithms Overview Five algorithms used in the GeneMapper software are discussed in this appendix Peak Detection uses Basic Advanced or Classic mode to detect peaks and process data Size matching calling matches found peaks to size standards Binning determines bin centers for genotyping Allele calling produces a consensus call based on several allele calling algorithms Quality value determination assigns a quality value to size and allele calling algorithms See Appendix A
97. andard 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 Using the Size Standard Settings 10 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description 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 10 4 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Creating Editing a Size Standard Overview To create a size standard you must first specify the parameters under which the size standard is created and then define the peaks using the Size Standard Editor When editing a pre defined size standard only the Size Standard Editor dialog box will be shown Specifying Size To specify size standard parameters Action Select Tools gt
98. anel data is as follows Data 4 Kit chemistry kit or product Panel Marker Bin A kit is a collection of panels A panel is a collection of markers Bin sets are collections of the expected allele locations for markers contained within a kit A different bin set should be used for each instrument type as variations in run conditions and polymers affect allele positions The GeneMapper software requires kit panel and bin set names to be unique throughout the database and requires marker names to be unique within a panel The Panel Manager enforces these requirements during user entry and editing of names as well as during data import Using Panel Manager 4 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 marker has a comment field Panel Manager The toolbar icons enable specific Panel Manager actions similar to their Toolbar associated menu items as described throughout this chapter Position your mouse over an icon to view its tooltip description 12 3 45 6 7 8 i xX IBI oe I Lae Bin Set SNP Demobins Dd Enta m a a fa Item Description 1 New Kit Panel Marker icon automatically updates to reflect the current action dependin
99. anes t em Bi WEN el it Fale ME Y Sample File Sample Name Pand os sQ DEE freser se 01_Pana o1 m ho ia Dm j 262 266 270 274 278 282 286 290 294 298 302 306 310 314 318 322 326 330 334 338 342 36 350 1040 360 880 800 720 640 560 480 400 320 240 160 30 okee Using Plot Windows Samples and Genotypes 7 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Allele Shows hides label editing effects Changes Turned Off Deleted label disappears Edited Added label looks normal 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 Label changed Allele label is shown with a double upper bar Charged al 152 Label added Allele label is shown with a double upper bar Added al 52 Controls To Controls whether or not control samples are displayed at Top 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 7 16 Using P
100. anteed 888 88 SOURCE www artisantg com Classic Peak Detection Algorithm This algorithm is similar to the GeneScan software on the MacIntosh operating system method of analyzing size standards and performing peak detection Refer to Appendix B Software Genotyping Algorithms for additional information Analysis Method Editor Microsatellite No Peak Correction E foi Using Analysis Methods 5 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Quality Tab The Peak Quality page sets the thresholds for warning flags in the Genotypes view in the Project window The PQV system uses these values to determine what types of warning flags should be set for each parameter When you click the Peak Quality tab the following page opens General Allele Peak Detector Peak Quality Quality Flags Signal level Homozygous min peak height fso0 0 Heterozygous min peak height fi 00 0 Heterozygote balance Min peak height ratio fos Peak morphology Max peak width basepairs fi 5 Pull up peak Pull up ratio pi Allele number j Max expected alleles k Factory Defaults OK Cancel The settings include the following 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 si
101. 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 and is set by the user to allow multiplexing of markers in a sample an ASR should be changed only when a marker is under development ASR boundaries can be edited in the marker table or in the Bin 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 Bin 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 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 first and then click Apply Then you can 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 Using Panel Manager 4 39 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Automatic Bin This section discusses the features of the A
102. are 2 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 containing 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 In Addition You can place key words in the Sample Comment or Color Comment field that distinguish samples from each other 2 10 Using GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneMapper Manager Introduction The GeneMapper Manager is a centralized interface within the GeneMapper software v3 0 for managing and organizing projects The GeneMapper Manager window contains six tabs that point to the following views Projects see Chapter 3 Using the Project Window Analysis Methods Chapter 5 Using Analysis Methods Table Settings Chapter 6 Using Table Settings Plot Settings Chapter 7 Using Plot Windows Sam
103. artisantg com The following table lists two common causes of bad matrices continued Problem Cause What to do Gel File rg 100 120 140 160 180 200 220 240 260 280 300 320 340 360 380 6B 11 test 2 test 2 EY 11 test 2 test 2 Dye Sample Minutes Peak Height Peak Area Data Point Peak 9 20 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Size Standard Settings Chapter Overview Introduction This chapter describes the Size Standard and Size Match Editors in the ABI Prism GeneMapper Software Version 3 0 In This Chapter This chapter contains the following topics Topic See Page About Size Standards 10 2 Size Standards Tab 10 3 Creating Editing a Size Standard 10 5 Size Match Editor 10 12 Size Calling Curve 10 18 Performing a Sizing Only Application 10 19 Using the Size Standard Settings 10 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About Size Standards What are Size Size standards are specific DNA fragments of known sizes After Standards defining the sizes of a size standard the GeneMapper software matches this definition to the internal size standard included with the run The software assigns the defined size values to
104. asic peak detection algorithm is used for most applications Peake Detector Peak Quality Quality Flags Peak Detection Algorithm Basic 7 Minimum Peak Height General Allele C Automatic User specified rfu Factory Defaults OK Cancel The application sets the level automatically for these five dye colors when Automatic is selected 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 Selecting the User Specified option button enables the number entry fields for five dye colors These numbers are the minimum signal strength that will be identified as a peak in relative fluorescent units For all dyes the default is 50 minimum is 1 and the maximum is any number of 10 digits Using Analysis Methods 5 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Advanced Peak Detection Algorithm This algorithm is similar to the ABI PRISM GeneScan software on Windows NT operating system method of analyzing size standards and performing peak detection Information on the individual parameters can be found in Appendix B Software Genotyping Algorithms Analysis Method Editor Microsatellite 5 12 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guar
105. ata and bin set data These files are tab delimited and the following rules apply 4 32 Files to be imported must have the correct number of columns in the correct order On import panel and bin set names are checked for uniqueness in the GeneMapper 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 If any errors are found during import no data from the file is accepted by the Panel Manager Lines beginning with are comments and are ignored on import Both Panel and Bin set file formats have a header line with a GeneMapper software version string Version GM v 3 0 If the first line of a file to be imported does not begin with Version GM then GeneMapper software handles the import as a GeneMapper software 1 x panel import If the file is in GeneMapper software 1 x format then it is imported without errors If the file is in GeneMapper 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 GMoistribut
106. ated list of allele names in the allelic Alleles ladder No validation Applicable only to Human Identification HID genotyping or applications that use allelic ladders 4 36 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Bin View for Microsatellites Only Introduction The Bin view enables you to view create and edit bins The data displayed in the Bin view imported into the Panel Manager using the Import BinSet command is used to create the bins associated with the marker Bins allow the user to average the minor variations in size that occur from run to run and capillary to capillary Panel Manager x File Edit Bins View Sx 8 Mee JAD Bin set OEG H Deaeoe m mlm la PEO D152797 Bins 102 104 106 108 110 112 114 116 118 120 122 124 126 128 130132 134 136 138 140 D182800 07 D18234 D18249 018450 018196 D18255 D1S2667 0182785 0182890 D18484 D18213 D1S2878 02 D15208 6a pier Z oot AMAMA n n 2 55 2 H 350 982 101 4 1046 1078 1110 1142 117 4 1206 1238 1270 1302 133 4 136 6 1398 1430 1452 amp Reference Samples Location 121 98 bp OK Cancel Apply Description of the Reference Data Bins and Allele Calls Bin View Reference data includes descriptions of bins name location and boundaries and the sample allele calls used to construct the bins In the Bin view the vertical bars are b
107. athematical formula used is AJLA to oY a AQ AQ X AQ MQ GQ MQ x PQV x POV x xPQVj a AQ Allele Quality is a function of quality value assignments for the following sizing quality allele calling quality bin assignment quality and bin quality GR2119 b MQ Marker Quality is modified by user defined PQVSs to generate the final GQ value PQVSs are weighted from 0 to 1 The actual value of a PQV in the equation is 1 minus the weight 0 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 failed Between 0 and 1 the higher the value the greater the impact on GQ PQV filtering is controlled by the threshold set by GeneMapper software users and remains fully functional irrespective of what weight is chosen Process Quality Values A 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Software Genotyping Algorithms Appendix Overview Introduction This appendix provides a description of the new genotyping algorithms used in the ABI PRisM GeneMapper Software version 3 0 and a detailed description of the Peak Detection Basic Classic
108. aximum range panel size range most recent Scale To range X Axis Scale for SNP data bi ca Scale to maximum Y Axis Scale gt Scale to panel size range Dyes b 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 Scale to maximum The electropherogram being viewed is scaled to its maximum base pair length as collected from the instrument Scale to panel size range The electropherogram being viewed is scaled to the size range available for the selected panel 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 Click and drag with the left mouse button the desired region on the X axis OR Right click on the X axis to open the X Axis Scale menu command options 4 20 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com X Axis for Microsatellite data x cale scale to tannin Y Axis Scale gt Scale to marker size range Dyes gt 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 Scale to maximum The electropherogram being viewed is scaled to its maximum base pair length as collected from the instrument Scale to
109. be checked To use the Invalidate Genotype Quality feature in the sample plot Step Action 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 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 Invalidate Genotype Quality feature in the genotype plot Step Action 1 Select the sample header of a plot to select it 2 Right click on the header to open the Override Genotype Quality dialog box 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 Using Plot Windows Samples and Genotypes 7 31 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Plot Settings Editor Chapter Overview Introduction This chapter describes how to use the Plot Settings tab in the GeneMapper Manager window of the ABI Prism GeneMapper Software Version 3 0 and describes how to use the features of the Plot Settings Editor In This Chapter This chapter contains the following topics Topic See Page Plot Settings Tab 8 2
110. below For more information see Appendix C Project Window Software Interface GeneMapper Project Microsatellite Project 1 File Edit Analysis View Tools Help 2 Ka amp BS a Ahh LLL EWI re Table Setting Applied Biosystems Appili z m 3 Project Samples Genotnes 3 7 Status Sample File Sample Name Comments Sample Type Analysis Method Panel _ Si 01_10030 6 ie 01_100301fsa 100301 None Sample microsatellite Default None G 4 02_1004 7 i 02_100405 fsa 100405 None Sample microsatellite Default None G 5 et is i 03_100501 fsa 100501 None Sample microsatellite Default None G 05 to004 9 i 04_100302fsa 100302 None Sample microsatellite Default None G os_10050 10 05_100406 fsa 100406 None Sample Microsatellite Default None G 071003 06_100502 fsa 100502 None Sample microsatellite Default None G 12 i 07_100305 fsa 100305 None Sample Microsatellite Default None G Progress Status a Ee E 6 The following table describes the parts of the main window in the above figure Parts of the main window Item Name Description 1 Drop down These menus are described in Project Window menus Menus on page C 9 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 Samples Genotypes View on page C 2 and Genot
111. 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 lg View BelectAl Add Samples to List Ctrl D Sear Clear All gt GFA Delete Options Add Sample Files Edit Menu Commands Item Description Enabling Select All Selects all the items of a given Enabled when a single Ctrl A type valid selection has been made e g sample file Project Window Software Interface C 37 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Add Sample Files Edit Menu Commands continued Item Description Enabling Add Samples to List Ctrl D Adds the selected item s from the Files GeneMapper database or Sequence Collector to the Samples to Add list Enabled when a sample file or folder with files in the Files GeneMapper database or Sequence Collector is selected Clear Delete Clears deletes the selected item s in the Samples to Add list Enabled when a valid selection is made Clear All Hierarchical menu Clears all sample files and folders from the specified field Samples to add Criteria Note The Criteria subcommand functions
112. by the appropriate filtering method N A indicates that no comments can be made for these rows Filtering Properties Summary For more specific information see Filtering Controls on page 6 10 The filtering properties for a Project table column are controlled in the Filtering and Content columns Modifying these settings updates the property in the Column Filter table For more information see Filtering Controls on page 6 10 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 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 Using Table Settings 6 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Allele Settings The Allele Settings box on the Genotypes page of the Table Setting Editor window controls how the Allele Size Height Area Mutation and Comment columns are displayed in the Genotypes view Allele Settings Number of Alleles k Keep Allele Size Height Area Mutation and Comment together Cancel Allele Settings Description Number of Alleles The number of alleles for which genotypes are displayed The default value is 2 This value
113. chnology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Add Samples Tab Elements Element Description Set Analysis Method to drop down menu and Read from the Sample option button When you add samples the Analysis Method property is set to the value in the text box which contains the list of analysis methods from the GeneMapper database Set Size Standard to drop down menu and Read from the Sample option button When you add samples set the size standard property to the value in the text box which contains the list of size standards from the GeneMapper database 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 Select a Panel for all samples option button and window 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 choices may be made 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 The Select a Panel window contains the list of panels from the GeneMapper database Set Panel to H A LM o Select a Panel for all samples l3 E Arr C Read from the Sample Sheet Sa
114. cifying panels Table for defining markers within a panel Graphical editor for creating and editing bins Data for the panels is stored in the database Clicking the Apply button after making changes to a panel sends all changes to the database but keeps the window open for further edits 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 4 2 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com When to Click the After deleting a kit panel marker or bin set Apply Button After making a number of edits After creating a bin set Displaying the To display the Panel Manager window either Panel Manager 4 Click W Panel Manager from the Project window toolbar Window or Select Tools gt Panel Manager in the Project window File Edit Bins View oR ee I inset 7 Reece PE E Panel Manager Kit Nane Kit Type Eana H EJSNPdemo SNPdemo SNP none m ELMS MD10 LMS MD10 Microsatellite none JAmpFISTR_Panels_v3 i ELMS HD5 AmpFISTR_Panels_v3 Microsatellite none 3 LMSHD5 V2 5 LMS HD5 Microsatellite none H ELMS LD20 LMSHD5 V2 5 Microsatellite none E EJLMSMD10 V2 5 LMS LD20 Microsatellite none LMSMD10 V2 5 Microsatellite none amp Reference Samples OK Cancel Apply Hierarchy of Panel Conceptually the hierarchy of p
115. ck A The second type the Sizing Quality SQ parameter reports results as Pass I Check A or Low Quality On the Genotypes page The SHP AN AE OBA SPA Bin PHR LPH SPU BD SP OS CC OVL NB and DP parameters report results as Pass B or Check The second type the Genotype Quality GQ parameter reports results as Pass B Check A or Low Quality Note On the Genotypes page the ADO and AE parameters are also PQV parameters but they report results as checks gt lt instead of colored flags A Low Quality red octagon quality value for the Sample SQ indicates that the sizing standard has failed A Low Quality value for the GQ parameter means the analysis has failed for that marker You can set the thresholds for the SQ and GQ values in the Analysis Method Editor on the Quality Flags page Process Quality Values A 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com SO l JOPPe 10 JON ALATA ATAJA AJA ATAATA A ALA dNS gt ALATASA ALATA ATATA TSA A A AJA aH Z a A AIALA ALA A A AIAILA A A A aon epyoejonu p Al AIALA ALATA ATAT ATATAJ ALA ALA LALA WI OS ANSS 4SNW SNSS dd 9N 1A0 00 SO dS Gd NdS Hd1 YHd Nid VdS VEO NV dHS AV OdV anieA AOd aiqeoijdde jon yew YOOUD ON sishjeue siy YIM peasy A el
116. ck this box for multicapillary instruments This user function was performed in Data Collection Used to specify that the GeneMapper 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 them see Chapter 9 Creating a Matrix File Smoothing option buttons Used to help reduce the number of false peaks detected by the GeneMapper 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 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Size Calling Method Parameter Options Click a radio button to select the desired size calling method The GeneMapper software uses these methods to determine the molecular length of an unknown fragment Description of the size calling method parameter optio
117. cleotide only analysis Not editable Process Quality Values A 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description AN Allele Number A flag Pass J 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 Triggering this flag reduces the final PQV Genotype value GQ parameter to zero O multiplier Not editable OBA One Basepair Allele 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 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 This flag is used only for LMS Microsatellite dinucleotide markers only Not editable SPA Single Peak Artifact A flag Pass J or Check A is automatically displayed when the marker signal contains single peaks due to some problem in electrophoresis The flag is triggered when there are no peaks present within a two base pair range prior to an allele peak This feature detects the absence of stutter peaks which indicates nonmicrosatellite peaks This flag is used only for LMS Microsatellite
118. columns 3 18 Samples View column filtering settings 6 10 Save Project Save Project As dialog boxes C 30 Sequence Collector database tab window elements C 44 tab view elements C 21 Sequence Collector tab C 44 size calling curve 10 18 black curve 10 18 red curve 10 18 size calling method Cubic Spline Interpolation B 34 Global Southern B 37 Local Southern B 35 size calling methods classic and advanced B 32 Cubic Spline B 34 Least Square B 32 Size Match Editor Edit Menu 10 14 procedure for editing a peak 10 16 View Menu 10 15 when to use 10 12 Size Match Editor window 10 12 editor commands 10 13 elements 10 13 toolbar 10 16 using 10 16 size standard 350 All size standard D 3 400 HD D 4 500 All D 6 creating and editing 10 5 specifying size standard parameters 10 5 Size Standard Editor window using in basic or advanced mode 10 8 using in classic mode 10 10 Size Standard tab 10 3 Size Standards advantages of using 10 2 what are 10 2 sizing only application sizing microsatellite reference data 10 19 sizing SNaPshot reference data 10 20 sizing only application 10 19 slope threshold B 30 peak end parameters B 28 Index 5 Guaranteed 888 88 SOURCE www artisantg com peak start B 28 Sort Dialog box C 47 SQ and GQ quality indications A 5 SSCP Minimum Peak Half width setting B 12 B 17 Stutter Distance 5 9 Stutter Ratio 5 9 T Table Manager content column dialog box 6 8 filtering controls 6 10 Table Setting Editor
119. creased from 15 red to 31 black data points Window Size 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 i Window size value of 15 red Electropherogram showing the same peaks as in the figure above see page B 21 after increasing the window size value while keeping the polynomial degree the same B 22 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Optimizing Peak Detection Sensitivity Example 1 Initial Electropherogram The figure below shows two resolved alleles of known fragment lengths that differ by one nucleotide detected as a single peak The analysis was performed using a polynomial degree of 3 and a peak window size of 19 data points R so sso goo szo aiso i90 4200 4210 4220 4230 4240 4250 4200 4270 4280 4200 400 4310 4320 4390 4340 4960 480 470 480 400 4400 4410 4420 zo tr ki t ir Eo 1 f aes A DyerSample Minutes Size Peak Height Peak Area Data Point Peak 206 29 1502 186 78 71983 24730 4342 Electropherogram showing two resolved alleles detected as a single peak
120. ction Default Analysis Type Bin Set Algorithm Microsatellite Microsatellite None Basic SNP Genotyping SNP Genotyping None Basic 3730 DS 33 Microsatellite 3730 DS 33 Basic For use Install Install Bins with the Applied Biosystems 3730 DNA Analyzer only Note These profiles can be altered but should not be deleted They are intended to be used as models for customizing new analysis methods 5 2 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About the Analysis The Analysis Methods tab contains a list of analysis methods in the Methods Tab database This tab is used to manage the contents of the project window To display the Analysis Methods tab select Tools gt GeneMapper Manager gt Analysis Methods iy GeneMapper Manager Name ___ Last Saved lower instrument Anatsis Tyne o a Default 2002 04 01 18 10 10 0 6M Microsatellite ER Se ee ny pa Parts of the Analysis Methods tab Item Description Name column Analysis method name Last Saved column Date Time stamp showing when the analysis method was last saved Owner column User name of the person who created the analysis method Instrument column The type of instrument used Analysis Type column Identifies the analysis mode of the analysis method i e HID Microsatellite SNaPshot Using Analysis Methods 5 3 Artisan Technolog
121. ction parameter options Item Description No Peak Correction No correction for doublets GS2500 Peak Makes the following peak size assignments for Correction GeneScan 2500 The right peak for all fragments 222 233 238 286 and 490 The left peak for all other splits Left Most Peak Chooses the left peak for every doublet Correction B 12 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Description of the split peak correction parameter options continued Item Description Right Most Peak Chooses the right peak for every doublet Correction Correction Limit Set a correction limit if correcting for doublets Set this value slightly 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 Software Genotyping Algorithms B 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Advanced Mode Peak Detection Algorithm Settings Overview In Advanced mode there are four main analysis parameter options They are as follows Ranges Smoothing and Baselining Size Calling Method Peak Detection Ranges Options The following are the Analysis Range options Item Description Full
122. ctor Field column Text field that contains the properties columns in the selected group criterion Condition column Field that contains conditions that the system will use to search for samples Search rules include Starts With Contains Equals Date criteria Presents a calendar widget to specify the date Criteria column Field for specifying the text string associated with the search rules which are case sensitive C 44 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sequence Collector Tab Elements continued Element Description Rules for Search Criteria Criteria accumulate as they are entered by the user 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 Search Criteria Cancel button Enabled when a search is in progress the search stops when clicked Note May not stop the search during data transfer from the GeneMapper database Search button Searches the database using the criteria list and displays the search results in the Search Results field Search Results Displays the results of the database search sample
123. ctropherogram but are not analyzed and do not display in the tabular data Software Genotyping Algorithms B 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Description of the detection parameter options continued Item Description For example Minimum Peak Half Defines what If this number is large Width constitutes a peak the software ignores i i ikes Use to specify the RA SEES 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 from Experiment with this 2 99 value to determine the for th A typical number might Bes ibiabet taane f data be 3 for microsatellites or 10 for SSCPs Split Peak Under denaturing conditions certain fragments in the GeneScan 2500 Correction standard display as doublets or split peaks This standard has labels Parameter Options 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 Description of the split peak corre
124. d 10 Matrix Cancel 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 If this occurs 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 Samples view filter settings Column Name Filter Settings Status Sample File Analyzed or Not Analyzed Show All Rec w Show All Records Analyzed Not Analyzed 6 10 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Samples view filter settings continued Column Name Filter Settings 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 Records Show All Records Show Records Containing Note This setting allows specification by a string Input this information into the Content column Sample Type Sample Control Allelic Ladder Primer Focus or Negative Control Sample Type Sample Control Allelic Ladder Negative Control Reference Data O
125. d in the preview below limiters E Treat consecutive delimiters as one M Tab I Semicolon T Comma I Space I Other E Text Qualifier z a preview Status Bample File 5 Sample Type An 01_100301 fsa Click the Next button to continue to Step 3 of 3 in the Text Import Wizard window Text Import Wizard Step 3 of 3 21x This screen lets you select each column and set olumn data format the Data Format General General converts numeric values to numbers date C Text values to dates and all remaining values to text C pate mov z Do not import column skip Advanced 1_100301 fsa N2_100405 fsa 03_100501 fsa 4 100302 fsa Cancel lt Back Next gt C As you can see the first column of the table is already selected Using the Project Window 3 21 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To open and save an exported text file in an Excel Step Action 5 Open the table file by performing the following actions a Hold down the Shift key Scroll to the last column of the table Click the last column of the table and release the Shift key Click the Text option button Click the Finish button to open the table file with all columns in text format ooo 6 Save the text file as a Microsoft Excel Workbook file The next time you open the file in Excel the t
126. deleting a table profile 6 9 editing a table profile 6 9 filtering controls 6 10 filtering settings genotypes view 6 12 filtering settings samples view 6 10 new table profiles 6 9 Table Settings Editor 6 4 General tab 6 4 Genotypes tab 6 6 Sample and Genotypes tab elements 6 7 allele settings 6 8 Samples tab 6 5 Table Settings tab overview 6 2 profiles 6 2 Technical support for computers with altered configuration F 2 technical support 1 12 Thresholds 5 16 troubleshooting bad matrix files causes 9 19 to 9 20 U User manual organization 2 12 Users tab elements C 24 WwW Warranty exceptions F 4 for computers with altered configuration F 2 Index 6 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com A rtisan Artisan Technology Group is your source for quality TecmoogyGroup new and certified used pre owned equipment FAST SHIPPING AND SERVICE CENTER REPAIRS WE BUY USED EQUIPMENT DELIVERY Experienced engineers and technicians on staff Sell your excess underutilized and idle used equipment TENS OF THOUSANDS OF at our full service in house repair center We also offer credit for buy backs and trade ins IN STOCK ITEMS www artisantg com WeBuyEquipment 7 EQUIPMENT DEMOS HUNDREDS OF InstraV ea REMOTE INSPECTION LOOKING FOR MORE INFORMATION MANUFACTURERS Remotely inspect equipment before purchasing with Visit us on the web at www artisantg com 7 for more o
127. dinucleotide markers only Not editable A 8 Process Quality Values Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description Bin Out of bin allele A flag Pass J or Check A is automatically displayed when the called alleles peak apex is out of the bin boundary Triggering this flag reduces the final PQV Genotype value The default is 20 0 2 multiplier of what it would be otherwise This flag is used for LMS analysis For HID this is labeled OL off ladder alleles Not editable PHR Peak Height Ratio A flag Pass J or 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 If there are more than two alleles present the calculation iterates through all the peak pairs PHR can be set in the Peak Quality tab of the analysis method Analysis Manager The default is 50 For LMS markers the ratio is calculated based on peak heights of the called allele peaks 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 Not editable LPH Low Peak Height A flag Pass I or Check A is automat
128. documents Download PDF documents Obtain information about customer training Download software updates and patches In addition the services and support page provides worldwide telephone and fax numbers to contact Applied Biosystems Technical Support and Sales facilities 1 12 About GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using GeneMapper Software Chapter Overview Introduction This chapter describes techniques for collecting and preparing sample data that will help you improve the overall quality of the fragment analysis sample files you import into the ABI PRISM GeneMapper Software Version 3 0 It also describes how to plan for use of the GeneMapper software features in your genotyping application In This Chapter This chapter contains the following topics Topic See Page Steps Required to Use GeneMapper Software 2 2 Installing and Registering the GeneMapper Software 2 3 Logging on to and Logging out of the GeneMapper Software 2 5 Importing Data from the Analysis Sample Sheet 2 8 GeneMapper Manager 2 11 Overview of the User s Manual 2 12 Using GeneMapper Software 2 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Steps Required to Use GeneMapper Software Using The following steps are required to use the GeneMapper software You GeneMapper
129. e Text Wizard has determined that your data is Delimited If this is correct choose Next or choose the data type that best describes your data riginal data type ile type that best describes your data Characters such as commas or tabs separate each field Fields are aligned in columns with spaces between each field Start import at row fi sj File origin windows ANSI z Preview of file K GM 3 0 pictures Project Folder exported table 1 bratusOSample FileOSample NameOCommentsOSample TypeOD nalysis Me 2 lta1seno1_100301 f sa01003010NoneDSampleODefaultOGreen_I_w3065400 a rue002_100405 fsa010040S50None0SampleODe faultONoneONoneOPorkcho 4 rueN03_100501 fsa01005010NoneDSampleODe faultONoneDGS400HDOPork s ruell04_100302 fsa01003020None0Sanple0De faultONonellS400HDOPork w a a Cancel lt Back n gt Finish 3 20 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To open and save an exported text file in an Excel Step Action 3 Move through the steps to import text a Click the Next button to proceed to Step 2 of 3 in the Text Import Wizard window b Select either Tab or Comma Delimiters according to how you exported the table if necessary Text Import Wizard Step 2 of 3 x This screen lets you set the delimiters your data contains You can see how your text is affecte
130. e components your computer system Software requires to run GeneMapper software Requirements IMPORTANT The Oracle database has an embedded license which only permits five named users Note In order to perform all functions in GeneMapper software with its embedded Oracle database please ensure there are at least 500 MB of free space on the partition where the GeneMapper database is installed It is possible to export your projects to a different drive or partition and then delete those projects from the database to make room for new projects See Chapter 3 Using the Project Window System Component Minimum Requirement Recommended Computer Intel Pentium III processor minimum of 540 MHz 256 MB of RAMa 6 GB hard disk free space Intel Pentium Ill processor faster than 700 MHz 512 MB RAM 10 GB EIDE hard drive 20 48X IDE CD ROM 10 100 NIC with RWV internal gt e Monitor 800 x 600 pixels size 17 inch monitor Larger monitor System Operating Microsoft Windows NT version 4 0 Service Pack 5 Microsoft Windows NT 4 0 Service Pack 5 or Windows 2000 Professional Service Pack 2 Ethernet Capability Network card for Oracle installation TCP IP must be installed prior to Oracle installation a There are functions within the GeneMapper software that run faster with more than 256 MB RAM Applied Biosystems recommends using at
131. e data electropherograms in the Panel Manager Plot view Checked when active Multiple dye colors may be selected Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Files like these 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 24 Bins Definition Example 4 26 Panel Definitions for SNaPshot 4 28 Bins Definitions for SNaPshot 4 30 Import and Export Files 4 32 Using Panel Manager 4 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Definition The examples below show panels defined in the format used by Example GeneMapper software v3 0 for a panel definition in a tab delimited text file txt using notepad or wordpad or a Microsoft Excel spreadsheet saved as a ixt file i Microsatellite Tutorial Panel txt Notepad E lol x File Edit Format Help MICROSATELLITE microsatellite Tutorial none none Blue 108 130 Blue 146 172 Blue 201 233 Blue 249 285 Blue 326 354 Green 104 138 Green 154 176 Green 201 245 Green 274 298 yellow 115 125 Yellow 167 yellow yellow Red Red Red Red Red NN 0 0 ie Q 0 0 0
132. e denoted by the red circles allele caller outputs are denoted by the 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 Examples of this occur in i and p Here the blue asterisks show the calls transmitted to the user Low quality values are reported because in both cases the first algorithm did not call and in i the black caller is not in agreement with the blue However despite these conditions the calls are correct The low quality values alert the user to potential problems such as the spurious peak in i and the high background in p Software Genotyping Algorithms B 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com a 1281659_27 b S1597_24 c 88258_24 d 125 5 1068 1354 f 196_24 g 158120 24 h 7S486_21 1118 241 488 1388 cast i 2382_20 j X81073_13 k 128310_7 1 118987_10 1011 1926 m 1271_14 n 1683103_6 0 118987_3 p 8S270_5 1426 B 6 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Quality Value The Quality Value Determination algorithm is used to create Process
133. ecks 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 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Import Bins Options x Select Option C Increase marker range to accomodate all bins C Delete all overlapping and out of bounds bins OK Cancel The Import Bins Options dialog box allows you to choose how GeneMapper software handles the bins being imported The numbered lines in the graphic above refer to the individual numbered descriptions in the figures on 1 The 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 Marker Range 100 110 Marker Range 98 114 Using Panel Manager 4 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The 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 A B c D E F G option2 gt Marker Range 88 116 Marker Range 100 110 The option shown below does not affect the marker allele size range all overlappi
134. ed 888 88 SOURCE www artisantg com Access GeneMapper Application Windows continued To see the Select And GeneMapper software displays the Size Standards Tools gt Currently Defined Size Standards GeneMapper x Manager Projects Analysis Methods Table Settings Plot Settings Matrices Name Last Saved Owner Type Description f n 1 GS400HD 2002 08 20 13 0 BasiciAd d Factory Provided Then select jom asic Advanced Factory Provide 2 65500 2002 08 20 13 0 gm Basic Advanced Factory Provided the Size 3 os500 250 2002 08 20 13 0 gm BasiciAdvanced Factory Provided 4 GSS00LIZ 2002 08 20 13 0 BasiciAd d Factory Provided Standards tab jam jasic Advanced Factory Provide 5 65500 250 UZ 2002 08 20 13 0 gm Basic Advanced Factory Provided 6 o8120LIZ 2002 08 20 13 0 gm BasiciAdvanced Factory Provided 7 377_G5_HID_GS500 2002 08 20 13 0 gm BasiciAdvanced Factory Provided 8 377_F_HID_GS500 2002 08 20 13 0 gm Basic Advanced Factory Provided 19 CE_G5_HID_GS500 2002 08 20 13 0 gm Basic Advanced Factory Provided 10 CE_F_HID_GS500 2002 08 20 13 0 gm BasiciAdvanced Factory Provided 11 GSS00LIZ_3730 2002 08 14 14 3 gm Basic Advanced 35bp 250bp and Size Match Analysis gt Size Match Editor Editor Size Match eE 2 Editor Edit View Tools xe or click the 20Samplet fsa Size Matches Size Calling Curve Icon Sizing Quality
135. eed sponse iget sige tea eee ce yes 6 2 Table Settings Editor 2 0 00 cece cece eee ee ete eee 6 4 Using Plot Windows Samples and Genotypes ChapterOverview oi sseswe ec etnies beet sides be beg pE E 7 1 About the Plot Window 0 0 0 eee eee eee 7 2 Plot Window Toolbars 0 0 00 eee cece ee eee 7 4 Plot Window Menus socn osie seh aceite gpk i ae ee goad hie bones 7 5 Using the Plot Settings Editor Chapter Overview o ssc ee cise bed ER aa RG dew bs da pha ds 8 1 Plot Settings Tab sspe ios riii a aii Maange e ae 8 2 Plot Settings Editor 0 eee eee ene neers 8 4 iv Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 9 Creating and Evaluating a Matrix Chapter Overview ees ees oyaeoe bo59e Vavde Coe ieee Bieehgeeawey cae 9 1 About the Matrices Tab 1 2 cece eee 9 2 About Matrics esie eke eka oye dba ee hae ee woe REA pias 9 3 Process of Creating a New Matrix 0 002 ceee eee eee eee 9 8 Choosing a Data Point Range for the Matrix Calculation 9 10 Generating a New Matrix 20 0 0 0 cee eee eee 9 12 Assigning the Matrix to Samples 0 00002 cee eee eee 9 16 Evaluating the Matrik 02 0625 ott benit age dengan debe ates earn ck oe 9 17 Using the Matrix with Data Collection 00 00005 9 18 Causes for Bad Matrices 0 0 0 00 eee eee 9 19 10 Using the Size Standard Settings Chapter OV
136. eight of the peaks that you want to detect Examining Peak To examine how GeneMapper software has defined a peak select Definitions View gt Show Peak Positions The peak positions including the beginning apex and end of each peak are tick marked in the electropherogram B 20 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Effects of Varying The figure below depicts peaks detected with a window size of 15 data the Polynomial points and a polynomial curve of degree 2 green 3 red and 4 Degree 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 2000 Polynomial curve of degree 4 1600 black Polynomial curve of degree 3 red 1000 Polynomial curve of degree 2 green 500 Electropherogram showing peaks detected with three different polynomial degrees Software Genotyping Algorithms B 21 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Effects of In the figure below both polynomial curves have a degree of 3 and the Increasing the window size value was in
137. ele Name User Name Modification Date Action and Comments Using Plot Windows Samples and Genotypes 7 27 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Rename Rename Allele 5 Allele v Allele Edit Comment Ctri O 6 Add Bin Cti B 7 Edit Bin Ctiu 8 Delete Bin Delete 9 10 11 12 13 14 15 2 Custom Allows the user to change the allele call on a labeled peak a Select Peak Selection mode b Select a labeled peak using the left mouse button c Select the Rename Allele menu item or click with the right mouse button to open the allele edit options and click Rename Allele d 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 the user to enter an allele comment for the selected peak The allele is then renamed to the selected allele call If Show type of edit prefix is turned on in the plot profile the word Changed displays inside the label box Allele Edit Controls whether or not the Allele Comment dialog box Comment opens when making label changes Checked when active cue To view the comments entered Show the AE Comment column in the Project Window Genotypes Table or Select the History option when right clicking a selected labeled peak or Double click a label 7 28 Using Plot Windows Samples and Genotypes Ar
138. elieved 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 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 Software has not undergone specific developmental validation for human identification applications Human identification laboratories analyzing single source or parentage samples which choose to use GeneMapper Software for data analysis should perform their own developmental validation studies Applied Biosystems ABI PRISM AmpF STR GeneScan Genotyper LIZ and SNaPshot are registered trademarks and the AB design Applera Expedite FAM GeneMapper GeneScan POP 6 ROX and TAMRA are trademarks of Applera Corporation or its subsidiaries in the U S and 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
139. en 35 and 500 base pairs The native fragments are uniformly spaced to provide accurate base calling The GeneScan 500 250 contains all of the fragments in the 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 Prepared The GeneScan 500 all Size Standard is prepared by Psi digestion of plasmid DNA followed by ligation of a LIZ or ROX 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 Lengths molecular lengths nucleotides for the 16 fragments Fragment Fragment Fragment Length Length Length 35 160 400 50 200 450 75 250 490 100 300 500 139 340 150 350 D 6 GeneScan Size Standards Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Electropherogram The screen below shows an electropherogram of GeneScan 500 run of GeneScan 500 under denaturing conditions Q Size Match Editor Edit View 2 Size Standard Size Match Quality 0 98 300 340350 250 200 150180 od 139 n a00 460 WAAL 4598 492 5228 5544 5860 6176 6492 6808 2000 i bs a
140. ength 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 wp Size Match Editor x Edit View Eg Size Standard Size Match Quality 0 99 3000 300 340 360 150 160 2m Ga 130 2000 100 iji l 2235 2465 2695 2925 3155 3385 3615 3845 4075 4305 4535 4765 4995 5225 5455 5685 595 6145 6375 6605 oK Cancel GeneScan Size Standards D 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneScan 400HD Size Standard About This Size Standard Special Uses How It Is Prepared Fragment Lengths Denaturing Electropherogram 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 contains 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 Leng
141. er table for that panel The Marker table shown below enables you to view create and edit marker names and associated data Panel Manager File Edit Bins View ae sew B E LMSMD10 V2 5 4 Panel02 MD10 V2 Ef Panel03 MD10 V2 Ef Panel04 MD10 Y2 Ef Panel05 MD10 V2 Ef Panel06 MD10 V2 Ef Panel07 MD10 V2 Ef Panel08 MD10 V2 Ef Panel09 MD10 V2 Ef Panel 0 MD10 V2 4 Panelt1 MD10 v2 Ef Panel 2 MD10 V2 EI Panel 3 MD10 V2 EI Panel 4 MD10 V2 EI Panel 5 MD10 V2 D15249 Blue Bin Set z 166 176 Herms ma EEE none D152800 Blue 207 207 D15234 Blue 270 274 D15450 Blue 331 339 D15255 Green 88 98 D152667 Green 137 141 0192785 Green 178 182 0192890 Green 210 214 D15484 Green 274 276 D15196 Green 326 326 D18213 Yellow 105 115 0182878 154 168 amp Reference Samples Yellow OK Cancel Apply Using Panel Manager 4 35 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Marker Table The figure on page 4 35 shows the columns in the microsatellite marker Columns table They are further described in the following table Marker table column descriptions Column Description Marker Editable Contains the name of the Marker Cell accepts Name
142. erview 20 00 05 eee beer ete tbe eee dew ce eb ase eee es 10 1 About Size Standards 00 0 eee eee 10 2 Size Standards Tab 2 2 eee ee ete tenes 10 3 Creating Editing a Size Standard 0 00 0 0000000008 10 5 Size Match Editor s c 6 2 che d ear dwanenls exe tvece deed dered toed ed ee 10 12 Size Calling Curve 0 0 0 cece eee ee erent eres 10 18 Performing a Sizing Only Application 0008 10 19 A Process Quality Values Appendix Overview sees secos deiae dere eE Ea Eea eee A 1 Overview of Process Component Based Quality Values PQV A 2 Using Process Component Based Quality Values A 3 B Software Genotyping Algorithms Appendix OvervieW 0 eect eee eee B 1 GeneMapper Software Genotyping Algorithms B 2 Basic Mode Peak Detection Algorithm Settings B 8 Classic Mode Peak Detection Algorithm Settings B 9 Advanced Mode Peak Detection Algorithm Settings B 14 v Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Detection Polynomial Degree and Peak Window Size Parameters B 19 Optimizing Peak Detection Sensitivity Example 1 B 23 Optimizing Peak Detection Sensitivity Example 2 B 25 Optimizing Peak Detection Sensitivity Example 3 B 27 Peak Detecti
143. es to Micosoft Windows Files Note Decompressing the CONVPROG HQx file creates a 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 To convert Macintosh computer sample files for use on a computer running Microsoft Windows operating system Step Action 1 Double click the Sample File Mac to Win to start the program Sample File Win to Mac The following dialog box opens AppleTest J el ual o bate raid 2 gt Qi Hard Disk Tool kit 12 17 98 z G parent Today db y Contains windows files Today b a rest of System stuff 12 11 99 b 2 Sequencing Analysis 3 4 1 5 17 00 db a System Folder 12 11 99 Where are the files to convert to Windows format New Cj J Cancel J Choose J 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 Sample File Conversion E 3 Artisan Tec
144. expands with the subordinate level indented Clicking the controller a second time will collapse the level back to its original form 3 Project Project Linkage __1 Run folder 01_100301 fsa 02_100405fsa 03_100501 fsa Sample files Artisan Technology Group Quality Instrumentation 04_100302 fsa 05_100406 fsa 06_100502 4sa 07_100305 4sa Using the Project Window 3 7 Guaranteed 888 88 SOURCE www artisantg com Once you have added files to the Project window the following rules apply 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 samples in the selected Run folder are displayed in the table select a Sample file in the Navigation pane sample information associated with the selected sample is displayed in the right pane See figure on page 3 8 click the Raw Data tab with the Info view displayed raw data associated with the selected sample is displayed in the right pane See figure on page 3 9 replacing the Info view display click the EPT Data tab sample information associated with voltage power current and run temperature is displayed See page 3 10 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
145. f the standard fragments Software Genotyping Algorithms B 35 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com B 36 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 val unknown ue Only the region of the size ladder near the fragment of 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 Step Action 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 A curve is then created by using three standard points two points below and one point above the fragment and a fragment size is determined Another curve is created by looking at an additional set of three points one point below and two points above the fragment and another value is assigned The two size values are averaged to determine the unknown fragment length Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Global Southern This method is similar to the Least Squares method in that it Method compensates for standard fragments that may
146. fecting all projects analyzed using this panel 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 Discard Changes This option allows you to discard any changes made to markers and bins Using Plot Windows Samples and Genotypes 7 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Command Description Print Automatically prints the contents of the plot window in the Ctrl P following order controls samples genotypes table if any Note Customize the view prior to printing Export Table Opens the Export Table dialog box allowing the user to Ctrl E export either the sizing or genotypes table shown in the Samples Plot window feptxport rabe Lookin 9 C GeneMappeniGMdistributionas mg E app _ Database docs Examples JRE a GeneMapper_Log txt a GeneMapper_Preferences X Show files of type Jal Files x Cancel Export file as Tab delimited text b 7 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 a
147. ff scale Sizing Quality Invalidated Yes or No 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 W Show All Records Show Pass Show Low Quality Using Table Settings 6 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 6 12 Using Table Settings Samples view filter settings continued Column Name Filter Settings Sizing Quality SQ Show Pass Show Check or Show Fail Show All Rec M 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 no instance of the type specified exists the Genotypes table will be displayed blank If this occurs return to the Table Manager and reset the filter 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 Genotypes view filter settings Column Name Filter Settings Sample File Sample Name Sample ID Run Name Panel Marker Allele Size Height Peak Area Mutation AE Comment Integration Comments User Defined Columns
148. files Columns in the table include Sample File plus others to match the fields specified in the search criteria Add To List button Adds the selected items from the Results field to the Samples To Add field Samples To Add field Contains the folders and files that the system will add 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 window Project Window Software Interface C 45 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Edit Menu Dialog Boxes Find Dialog Box The elements of the Find dialog box is described below Find what in column Sample File Name Project Window Find Elements 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 C 46 Project Window Software Interface Artisan
149. files to generate reports easily for downstream processing i e export text files to other applications For more details see Table Settings Editor on page 6 4 Analysis Method Editor GeneMapper Manager This page enables you to create and edit analysis methods including General properties such as type of analysis Microsatellite or SNP Genotyping Allele calling properties such as the bin set to be used and marker repeat information di tri or tetranucleotide type cut off value PlusA distance PlusA ratio stutter distance and stutter ratio etc Using the Project Window 3 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 3 4 Using the Project Window Peak Detector parameters and Algorithm type basic advanced or classic Marker Quality Values to specify various requirements for analysis Peak Quality parameters such as homozygous and heterozygous peak heights maximum expected alleles etc Quality flags settings and PQV threshold values pass and fail ranges for sizing and genotype quality values For more details see Analysis Method Editor on page 5 5 Panel Manager This window enables you to create and edit panels or reference data and has the following components A tool for creating product or chemistry kit folders A table for specifying panels A table for defining markers within a panel
150. for Microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot samples When you turn off a specific dye color for SNaPshot samples the bins for that color turn off also 7 14 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Bins Combine Dyes Mode continued gt Samples Plat BE Eloi File Edit View Tools Alleles Help a is i fee ey SNP Defaut z E Panes 2 z EE Eee Bo Man Eal MA af Me Sample File Sample Name Panel os sa 5_E03_09 fsa 5 plex Sampi J m l 17 19 21 23 25 27 29 31 33 35 37 3 MN B 45 g 649 1 53 55 57 59 6 amp 1 63 65 67 3220 _ i ae Bade mer Faer Ejer E heer 2862 2504 znas 1788 141 1073 715 357 h oO al A al GA lal Gj aT BLCJTI fac Ly es a Note Bins are enabled in Separate Dyes mode only for Microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot samples When you turn off a specific dye color for SNaPshot 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 nn atc File Edit View Tools Alleles Help wicrosatette 0 P
151. g B 4 Alleles tab range filter 5 10 SNP cutoff value for SNaPshot 5 10 Analysis Method Editor Alleles tab 5 7 General tab 5 7 Peak Detector tab 5 11 Peak Quality tab 5 14 Quality Flags tab 5 16 analysis methods 5 2 analysis parameters setting analysis range options B 14 data processing options B 15 Analysis Tab view elements C 22 Apply button 4 2 4 3 4 7 10 13 10 17 Apply To checkbox 7 11 Artisan Technology Group Quality Instrumentation Automatic bin assignment 4 40 builder 4 40 Automatic Bin Assignment ABAA 1 3 Automatic Bin Assignment Algorithms ABAA 1 4 Automatic Bin Builder ABB 1 3 1 4 B Bin Set alleles tab 5 8 choosinga 5 8 Bin view cursor location information 4 38 description 4 37 genotype quality symbols 4 38 Marker Allele Size Range ASR 4 39 purpose of 4 37 X and Y axes 4 38 Bins definition format of text file 4 26 Bins definitions for SNaPshot 4 30 C CD ROM contents 2 3 Collecting and preparing sample data techniques for 2 1 Command activation three ways of C 9 Computer configuration requirement F 2 technical support for altered configuration F 2 Cubic Spline Interpolation method B 34 customer support See technical support 1 12 Cut off Value 5 9 D Dye lanes window example of 3 14 Index 1 Guaranteed 888 88 SOURCE www artisantg com E Ease of use 1 4 EPT tab 3 10 Example Plot printout C 35 Example Table printout C 34 Export Project window 3 25 Export Table dia
152. g com Electropherogram After re analyzing with a value of 35 0 for the Slope Threshold for Peak After Adjustments 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 E oo zo zw ats zeo zwo zeo noo 200 amo pmo mo pao mso neo 2m0 m0 moo noo mo nw Electropherogram showing the effect of changing the slope threshold for peak end Software Genotyping Algorithms B 31 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Size Calling Methods for Classic and Advanced Modes Overview Least Square Method There are four size calling methods that you can use with the GeneMapper software v3 0 They are as follows Least Square Cubic Spline Interpolation Local Southern Global Southern Both Least Squares methods 2nd Order and 3rd Order use 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 is either a quadratic or a cubic function The software
153. g on the item selected in the navigation pane File menu 2 Clear icon performs kit panel marker deletion Edit x menu ie at 3 Icons select bin options Add Bin Delete Bin Edit Bin Bins menu rT 4 Icons allow viewing of Project alleles Bins menu ill 5 Icon switches view to full X Axis and Y Axis scales lt lt View menu 5730 08 33 instan Bi E 6 Pull down menu allows selection of a bin set for a selected kit 7 Icons select specific kit panel marker actions Bins menu m e E New Bin Set llr Add Reference Data Auto Bin E Panel Reference Data Auto Panel il a ja a fer 8 Dye color toggle icons View menu 4 4 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Manager The Panel Manager navigation pane is the primary navigation interface Navigation Pane 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 markers and bins The elements of this field are described below Element Description 3 Panel Manager The Panel Manager root node is the container for all panels It is the top level of the hierarchy 3 Panel Manager 9 LMS HD5 V2 5 1 JLMS MD10 2 5 Kits help users organize their panels in product or
154. ger 4 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Bins Menu The Bins menu contains commands to edit markers and bins EA View Edit SHE Marker StiHH Delete SNP Marker Str Shift D Add Bin Ctrl B Edit Bin Ctrl H Delete Bin Ctrl D New Bin Set Delete Bit Show Project Alleles Add Referer ji Panel Referer Auto Bin Auto Pane Citthee Edit SNP Marker The Edit SNP Marker command opens the Edit SNP Marker dialog box allowing the user to edit the marker name and bin information for the selected marker To enable this feature Select the menu item when a SNP marker is selected in the Panel Manager Plot tab OR Right click on a SNP marker or bin in the Panel Manager Plot tab Additionally SNP markers and bins can quickly be edited by changing the data in the Panel Manager Table tab for a selected SNP Panel 4 12 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Delete SNP Marker The Delete SNP Marker command deletes the SNP marker and associated bins selected in the Panel Manager Plot tab To enable this feature Select the menu item when a SNP marker or bin is selected in the Panel Manager Plot tab OR Right click on a SNP marker in the Panel Manager Plot tab Add Bin The Add Bin command creates a new bin associated with a panel as follow
155. gnal peak height setting set in the Peak Detector page will still be scored but the peak height flag will be triggered 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 value will be lower 5 14 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Morphology Max peak width basepairs default 1 5 Pull up peak Pull up percentage default 0 1 Considers the ratio of any color peak directly under a major peak Flags Spectral Pull Up quality value as in the Genotypes table Allele Number Max expected allele number default 2 If analyzing polyploid alleles enter in the maximum number of alleles you expect You must also set the table profile to display the correct number of alleles SNP for SNaPshot application only Double peak default 0 5 See PQV Flags on the Genotypes Page in Appendix A Using Analysis Methods 5 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Quality Flags Tab Use the Quality Flags tab to set the importance level of the PQVs available for microsatellite and SNP analysis The PQVs are
156. hat 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 7 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 7 4 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Plot Window Menus File Menu The File menu commands are used to perform the basic commands AES Edit View Tools Alleles H Save Panel Ctrl Shitt S Print Ctrl P Export Table Ctri E Close Plot Window Escape Command Description Save Panel Opens the Save Panel s dialog box informing the user of Ctrl Shift S what panel s have been modified and how many projects currently in the GeneMapper database will be effected The options are Panel Tutorial Panel 9 Was Modified Changes Will Effect 1 Project Save Panel C Save Panel As C Discard Changes OK E Save Panel This option allows you to save any marker and bin changes made to the current panel name af
157. he Info tab for the selected sample in the project Always enabled C 14 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window View Commands continued Ctrl Shift N Item Description Enabling Raw Data Switches to the Raw Data tab Always enabled Ctrl F2 for the selected sample in the project EPT Data Switches to the EPT tab for the Always enabled Ctrl F3 selected sample in the project Full View Shows the full view of the Raw Enabled only when Ctrl Data or EPT graph viewing the Raw Data or EPT Data of a sample Y Axis Scale Changes Y axis scale See Y Enabled only when Axis Scale on page C 15 viewing the Raw Data or EPT Data of a sample Show Switches the navigator pane Always enabled Navigator shows hides the pane Y Axis Scale The Y Axis Scale option is only enabled when viewing the raw data or EPT data of a sample file YA cale Scale to maximum Y Show Navigator Ctrl Shift N Scale to There are two options Scale to maximum Y scales to the maximum Y value for the selected sample Scale to opens a dialog box in which a user can define the value of the Y axis Enter Y Axis Scale Yalues x Minimum paua Maximum 5o00 cana Project Window Software Interface C 15 Artisan Technology Group Quality Instrumentation
158. he 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 Minimum Peak Height Automatic Minimum Peak Height User specified rfu Minimum Peak The Minimum Peak Height group box provides two parameter options Height Item Description Automatic option button This option button sets the level automatically for 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 option button This option button enables the number entry fields 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 8 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Pa
159. hing This algorithm uses a dynamic programming approach that is efficient Size Calling runs in low polynomial time and space and guarantees an optimal Algorithm 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 SizeSize Matching Algorithm Example An example of how the size matching calling algorithm works is 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 identifies all of the size standard peaks and removes the contamination peaks denoted by the black triangles from consideration 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 Bale mpate peaks found T T 200 4 3 o ire e Te f N N D ye N ie wo O q 8100 l l l vl v l y
160. hnology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Column Menus Columns are used to set up analyses after you have added samples to and Rules for a project Columns Column Menus Five columns in the Samples view of the Project window have drop down menus to apply settings to selected cells in the column Sample Type Analysis Method Panel Size Standard Matrix Note The Panel column displays a pop up window which contains a hierarchical list of kits and panels A panel is required for any allele calling analysis Each of these columns is used to apply needed analysis parameters to selected samples Different list selections can be made to the individual column rows representing samples Rules for Columns The following rules apply to column selections Clicking the header for a column selects the entire column Shift clicking selects a continuous selection range Ctrl clicking on individual cells makes discontinuous selections 3 18 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Applying a List Selection to an Entire Column To apply a Menu selection Step Action 1 Apply the selection to the top cell in a column a Select the top cell in a column This will display a drop down list of items b Select and click an item to apply the selection to the cell Se
161. hnology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To convert Macintosh computer sample files for use on a computer running Microsoft Windows operating system continued Step Action 3 Select the folder by single clicking its name 4 Click the Choose button at the bottom of the dialog box 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 Sample File Conversion Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Software Warranty Information Appendix Overview Introduction This appendix describes the software warranty provided by Applied Biosystems for the ABI PRISM GeneMapper Software Version 3 0 In This Appendix This appendix contains the following topics Topic See Page Computer Configuration F 2 Limited Product Warranty F 3 Software Warranty Information F 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Computer Configuration Configuration Applied Biosystems supplies or recommends certain configuratio
162. i 40500 5 2384 2700 3016 3332 T24 7a 7756 8072 8388 8704 e GeneScan Size Standards D 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sample File Conversion Appendix Overview Introduction This appendix describes how to use sample file conversion programs to prepare Macintosh computer generated fragment analysis sample files for transfer to a Microsoft Windows based format and vice versa In This Appendix This appendix contains the following topic Topic See Page Converting Macintosh Sample Files E 2 Sample File Conversion E 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Converting Macintosh Sample Files About Converting Applied Biosystems created two conversion programs that prepare Sample Files 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
163. ically 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 Process Quality Values A 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description SPU Spectral Pull Up A flag Pass J 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 LMS Microsatellite SNP and HID Not editable BD Broad Peak A flag Pass J or Check A is automatically displayed when the called alleles peak width is wider than a certain value The default of this value is 1 5 basepair This flag is used for SNP HID and LMS Microsatellite all markers Not editable SP Split Peak A flag Pass J or Check A is automatically displayed when a split peak is found b
164. ing Data from the Analysis Sample Sheet 2 8 GeneMapper Manager 0 0 eee ee eee 2 11 Overview of the User s Manual 0 00 00 e eee eee eee 2 12 Using the Project Window Chapter OVverviewW persie rrie cpta booed bees ede VaR e Eada ees 3 1 Central GeneMapper Software Window 0 02 eee eee 3 2 Details of the GeneMapper Project Window 000 3 5 Exported Project Tables 0 0 0 0 0c eee ec eee 3 20 Projects Page sjede sere o 5246652556024 aed OAR a E A E betas ee 3 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Panel Manager Chapter OVervieW 3 00 iy ceeeed baton Wade be dees bea EOE eg ves 4 1 Panel Manager Overview 0 cece eee teeta 4 2 Panel Manager Window Commands 0 0000 e eee eens 4 6 Formats of Panel and Bin Text Files 0 000 4 23 Panel Table VieW socwasac ce etdaes ee phate bs Fae edt AR ae 4 33 Marker Table View 0 0 reed ee Eee e a e s 4 35 Bin View for Microsatellites Only 0 00 rrna 4 37 Using Analysis Methods Chapter Overview serrie ocase draio upay eee bee ee ew eee eta 5 1 Analysis Methods Tab 0 0 00 5 2 Analysis Method Editor 0 0 00 eaa eee eee eee 5 5 Using Table Settings Chapter OVervieW 0 6 05 cae eh oad et barb beh ee eee eee 6 1 Table Settings Tab cccdisin deg
165. ings Editor Display Settings tab 8 9 General tab 8 4 Genotype Header tab 8 5 Labels tab 8 7 Sample Header tab 8 5 Sizing Table tab 8 6 plot settings editor 8 4 Plot Settings tab 8 2 Plot window Add Allele option 7 27 Add Bin option 7 29 Allele Changes option 7 16 Allele Edit Comment option 7 28 Alleles menu 7 25 bins 7 14 Controls To Top option 7 16 customizing 7 3 Delete Allele option 7 27 Delete Bin option 7 30 displaying 7 2 Dyes option 7 17 Edit Bin option 7 29 Edit menu 7 7 Editing Mode option 7 25 example of 3 14 Export Table dialog box 7 6 File menu 7 5 Full View option 7 9 Header option 7 21 Help menu 7 30 History option 7 27 labels 7 12 Marker Indicator option 7 21 Marker Margin option 7 22 Marker Range option 7 21 menus 7 5 Off scale Peak Indicator option 7 17 overview 7 2 Peak Positions indicators 7 14 Plot Settings option 7 24 Index 4 Plots option 7 18 PQV feature 7 31 Rename Allele option 7 28 Table Filter option 7 20 Tables option 7 18 7 20 toolbars 7 4 Tools menu 7 23 View menu 7 8 X Axis Scale option 7 10 Y Axis Scale option 7 11 PlusA Distance 5 9 PlusA Ratio 5 9 polynomial degree peak detection B 19 varying B 21 window size value B 22 PQV results A 5 Prefences Add Samples spellings and abbreviations C 20 Preferences Analysis tab C 22 Import tab C 18 Sequence Collector tab C 21 Users tab C 24 Print dialog box C 33 Process Component Quality Values PQV 1 4 Process Component Based Quality
166. ins 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 shown when those alleles are used in the autobinning process or in the manual process of bin creation Using Panel Manager 4 37 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Panel Manager x File Edit Bins View Bx iki Ik in Bin Set 31 00Avant D833 Tease Lia JPanel Manager GeneScan Insta 112 118 B 35 DS 33 1 0 D981690 D55644 D55424 D98288 D185462 D68289 D158117 a E9D08 30 E5 SNPdemo H E Splexdemo Eo mnj E Reference Samples DS33 LMS fsa 0 0 44 98 100 102 104 106 108 110 112 114 116 118 120 122 124 126 128 ee D20 119 4 v Lal pal 4 gt Marker 0208119 103 00 123 00 OK Cancel Apply If you analyze data using a bin set like that shown above you will see the s as shown above Then when you click the Show Project Alleles icon LI the s will be overlaid by a set of symbols marked in 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
167. ion Guaranteed 888 88 SOURCE www artisantg com Alleles Menu The Alleles menu is used to edit allele call labels on selected peaks and is only enabled when labels are displayed in the Plot window and one or more peaks are selected Help Editing Mode gt AG Aele Sine Delete Allele Delete History Rename Allele gt v Allele Edit Comment Ctrl 0 Add Bin Ctrl B Edit Bin Ctri U Delete Bin Delete 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 MANENE oil Binning Ctrl In Peak Selection mode Ctrl Peaks can be selected Allele calls can be edited 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 Continuous peak selection is available Use the Shift key or Click Drag inside the plot area Discontinuous peak selection is available 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 Using Plot Windows Samples and Genotypes 7 25 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Editing Mode In Binning mode Ctrl continued Marker si
168. ion app folder on your hard drive Panels and bin sets created in GeneMapper software v1 0 2 or GeneMapper software v2 0 are automatically updated when imported into GeneMapper software v3 0 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 called the Panel Table view Tasks you can perform in the Panel Table view include Editing panel names and entering comments in the panel table Creating new panels using the New Panel command Deleting an existing kit by selecting it in the Navigation pane and then choosing the Clear command Exporting panels and bin sets used for project analysis to backup Note Editing or deleting kits panels and bin sets will prevent your from displaying analyzed projects Export any panels and bin sets that you might use at a later date or for reanalysis of old projects gp Panel Manager x File Edit Bins View Xe Haa PW ensten gt ea he i Sg Panel Manager Kit Nene Et Type Comment H EJSNPdemo 1 SNP none 5 ELMS MD10 2 LMS MD10 Microsatellite none AmpFISTR_Panels_v3 C LMS HD5 3 AmpFIST
169. ions for each user To log on to the GeneMapper software Step Action 1 When you start the GeneMapper software application the following dialog box opens Select your user name if it exists enter your password and click OK BR Genetapper f A UserNamefon H Password Fep o Database Host cam75 AR aped ns 1999 2001 Applera Corp All Rights Reserved New Host Delete Host IMPORTANT If you do not have a User Name select the default gm enter the password ifa and click OK to launch the GeneMapper software You can go to Options from the Tools menu and create new users and passwords Using GeneMapper Software 2 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To log on to the GeneMapper software continued Step Action 2 Click the New Host button to open the New Host dialog box Enter new GeneMapper host information Host Name J Machine Type stand alone 7 j Cancel This option allows you to connect to GeneMapper software v3 0 databases on other computers 3 In the 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 Ifthe GeneMapper software cannot connect to the database host you entered the following error message opens You have entered an invalid host Click OK
170. iosystems 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 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 F 4 Software Warranty Informatio
171. is computer to the Data Collection computer Note GeneMapper software matrices are stored in the Oracle database Creating and Evaluating a Matrix 9 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Process of Creating a New Matrix Process Diagram The following diagram shows the procedure for making a new matrix Import samples into a new GeneMapper project ee View raw data to determine data point range for the matrix J Select Tools gt GeneMapper Manager f Select the Matrices tab J Click New _ Enter a matrix description J Select the number of dyes _ lt Select the sample file for each dye color J Insert the data point to start at for each dye color J Click Create es Review and edit the matrix result J Click OK to save and close the window Project window GeneMapper Manager window Matrix Maker window For sample preparation and loading information refer to the appropriate instrument user manual 9 8 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Steps to Create a The following table lists the steps to create a new matrix New Matrix Step Process See Page 1 Choosing a Data Point Range for the Matrix 9 1
172. 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 selected Ctrl F2 electropherogram See Chapter 3 Using the Project Window EPT Data Provides quick access to the EPT Data tab for the selected Ctrl F3 electropherogram See Chapter 3 Using the Project Window 7 8 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Full View Ctrl Automatically resets the X and Y axes of the electropherogram s currently viewed to their full scale size A File Edit View Tools Alleles Help Microsatenite i Panes 1 gt TE See Bia D ze a cal it fre Maja Sample File Sample Name Panel os sQ C006 _E019_Run_ E019 Tuterial Pan 480 182 184 186 188 190 192 194 196 198 200 202 204 206 208 210 212 214 216 218 220 222 224 i Zoomed In View La u F Note Double clicking with the left mouse button on an axis automatically resets that axis to full view i File Edit View Tools Alleles Help Microsatellite ji Panes j1 7 PE Sea Biz el Eak MA Yel M 4 Sample File Sample Name Panel os sa
173. is correct in most cases However when you are analyzing polyploid data you should change this value to the maximum expected allele number to view all calls 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 6 8 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Creating a New To create a New Table Setting Table Setting Step Action 1 Select GeneMapper Manager gt Table Settings and click New to open the Table Setting Editor window 2 In the General tab enter a name for the new setting and a description if necessary 3 Open the Samples and or Genotypes tabs and select the desired contents for the Samples and Genotypes tables in the corresponding tabs 4 Click OK to save the new setting Editing a Table To edit a Table Profile Setting Step Action 1 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 Set
174. ith the window size Use polynomials of degree 2 or 3 for well isolated peaks such as those from a size standard anda 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 For example 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 0 will move the extent of the peak toward its center B 18 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Detection Polynomial Degree and Peak Window Size Parameters About These Parameters How These Parameters 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 deg
175. ity Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window File Menu Commands continued Item Description Enabling Exit Alt F4 Exits the GeneMapper application Always enabled displays Save alert message if Project has pending changes Edit Menu The commands in the Edit menu are used to manage the contents of the Project window by performing standard actions like undo delete select and by enabling access to settings for Preferences ie Analysis View Tools Delete from Project Select All Ctri A Fill Down Ctrl D Find Ctrl F Sort Ctrl G Project Window Edit Menu Commands 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 selected message Deleting the selected sample s will delete both samples and results from project This action cannot be undone OK Cancel 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 Sequence Collector sample data Select All Selects all samples in the active Always enabled Ctrl A view Project Window Software Interface C 11 Artisan Technology Group Quality Ins
176. ization of the peak detection algorithm and quality value settings Se tine In the Peak Detector tab choose a peak detection Flowchart 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 Analysis Method Editor window Using Analysis Methods 5 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Accessing the Access the Analysis Method Editor in the following ways Analysis Method Click Tools gt GeneMapper Manager gt Analysis Methods Tab gt Editor New or Open Inthe Project Window click the Samples tab Click New Analysis Method to create a new method or double click an existing method to edit it Select a sample in the project window and click the Analysis Method Editor icon on the toolbar Edit the currently selected samples analysis method if necessary Note The Analysis Method Editor icon is not active unless a sample with an analysis method is selected New Analysis When you create a new analysis method the following dialog box Method Dialog opens prior to opening the analysis method editor This dialog box Box allows you to select the analysis type for your samples and controls the items available for customization within the Analysis Method Editor tab Select analysis type C HID C SNa
177. le you should only see blue sharp well defined singularly colored peak data Creating and Evaluating a Matrix 9 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Matrix with Data Collection Using Different Computers for Data Collection and Analysis Where to Store Exported Matrix Files Be sure to copy matrices generated and exported on the analysis computer the computer running GeneMapper software to the Data Collection computer if different Copying the matrices will ensure that the correct matrix name is stored in the sample file The proper matrix is required for accurate analysis of 310 and 377 sample files To transfer the matrix from one computer to another Step Action 1 Click the Matrices tab in the GeneMapper Manager window 2 Select Matrix to Export 3 Click Export 4 Select a name and location for the matrix 5 Transfer the mtx file to the data collection computer Store exported matrix files mtx intended for use by Data Collection software in 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 software matrices are stored in the Oracle database 9 18 Creating and Evaluating a Matrix A
178. lect all other cells in the column either as a continuous or discontinuous selection to which the list selection is to be applied Click Edit gt Fill Down to apply the choice made for the first cell to all selected cells Ctrl D Fill Down the Panel Column To fill down the panel column Step Action 1 Select the top cell in the Panel column to open the Select a Panel window Expand the appropriate folder Double click the appropriate panel Click Edit gt Fill Down Using the Project Window 3 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Exported Project Tables Opening Exported Project tables are exported as text txt files When they are opened in Project Tables in Microsoft Excel software use the Text Import Wizard as described in Microsoft Excel the following procedure so that all columns of the table are interpreted correctly To open and save an exported text file in an Excel spreadsheet Step Action 1 Start Excel software and either select File gt Open gt Files of Type or select Data gt Get External Data gt Import Text file Then select the txt file and click Import 2 Navigate to the folder containing your exported table files and double click the table file you wish to open The Text Import Wizard window opens Text Import Wizard Step 1 of 3 121 xi Th
179. led Genotypes View See page C 6 The tab for this view is enabled when results are present Samples that do not have a Panel attribute do not display in the Genotypes table Note Samples with no results do not display in the Genotypes tab Using the Project Window 3 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Samples View Microsatellite Default E E Project w EDatabasing Sample Type Analysis Method Panel Size Standard Matrix Run Name InstrumentType Instrument ID Run Date amp Time REF SQI SFNF MNF SNF 1 Control HID_Advanced Identifiler_v3 GS500UZ Databasing ABI3100 3100_1202_ 2001 03 16 11 41 2 Allelic Ladder HID_Advanced Identifiler_v3 GSS00LIZ Databasing ABI3100 3100_1202_ 2001 03 16 11 41 E E 3 Sample HID_Advanced Identifiler_v3 GSS500LIZ Databasing ABI3100 3100_1202_ 2001 03 16 11 41 nf E Genotypes View E GeneMapper Project SUSAN emon oven iso foorro fo e Note Shown with navigator pane closed Project Window The table below is provided as a reference for the Project window Toolbar toolbar commands Icon Description Creates a New Project fad Tooltip New Project 3 12 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Description Opens the Open Project dialog box Tooltip O
180. llector Project Window Software Interface C 21 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Analysis Tab The Analysis tab contains preferences associated with the analysis of data Startup Add Samples Sequence Collector Analysis Users Automatic Analysis I Automatically bring errors to the top of the table Quality Metrics Display Symbols C Numbers Mif only one labelled allele in a genotype then duplicate the label Duplicate homozygous alleles i Cancel Analysis Tab Elements Element Description Automatic Analysis When samples are analyzed the software sorts group box the table automatically to bring samples with errors to the top of the table Same functionality as Automatically brings the Bring Errors To Top icon in the Project window errors to the top of the table check box Quality Metrics Display Display quality values in project tables as symbols group box green square yellow triangle red octagon or Symbols button 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 Numbers button C 22 Project Window Software Interface Artisan Technology Group Quality Instrumentation Gua
181. log box C 31 F failed in lane size standards ability to correct 1 7 Filtering controls 6 10 Find command planning for use of 2 9 Find dialog box C 46 Fully automated operation 1 4 G GeneMapper access rules 2 7 accessing subordinate application windows 3 15 logging in and out 2 5 major steps in using 2 2 overview 1 2 registering 2 4 GeneMapper Manager Analysis Method Editor 5 5 Analysis Methods tab 5 2 Matrices tab 9 2 New Analysis Method dialog box 5 6 GeneMapper results exporting as text files 1 6 GeneMapper Software installing and registering 2 3 GeneMapper software hardware and software requirements 1 9 how it works figure 1 8 important features 1 6 read only access to the BioLIMS database 1 6 GeneMapper system dataflow 1 8 GeneScan Analysis Sample Sheet importing datafrom 2 8 Genotypes view Index 2 Artisan Technology Group Quality Instrumentation descriptions of columns C 7 Genotypes View filter setting 6 12 genotyping algorithms B 2 H High throughput 1 3 I Import Project window 3 27 Info tab 3 8 L Least Square method B 32 Local Southern method B 35 logging out of GeneMapper 2 7 M Marker 4 36 marker allele size range ASR 4 39 marker and bin definitions exporting 1 6 importing 1 6 Marker table columns 4 36 Marker Table view 4 35 matrices about 9 3 causes for bad 9 19 error messages 9 19 limitations 9 6 where to store 9 7 9 18 Matrices tab 9 2 matrix assigning to
182. lot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Off scale Peak Indicator Controls whether or not offscale peaks are highlighted in an electropherogram Checked when active When 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 La sox File Edit View Tools Alleles Help SNP Defaut gt fil Panes i z EE EEE Em MAN EAE m af a 4 Sample File Sample Name Panel 0S 8Q 3_co3 ossa 3 ie sampi a wt 28 3 7 S I S M ee 8 N S S a S e e eS ihe her fat Eper 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 av Blue Dye Ctrl 1 Plots gt v GreenDye Ctri 2 Tables b v Yellow Dye Ctri 3 Table Filter gt vRedDye Ctri 4 v Header v Orange Dye Ctrl 5 v Marker Range Ctri R v All Dyes Ctri 6 Using Plot Windows Samples and Genotypes 7 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description
183. ly 2 alleles GeneMapper software human identification and forensic applications a set of panels the grouping of panels in a kit is determined by the kit provider Linkage Mapping Set ABI chemistry using dinucleotide repeat microsatellite markers a known microsatellite or SNP 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 GeneMapper software project a collection of samples Primer extension based chemistry for SNP validation single nucleotide polymorphism used in this document to refer to SNaPshot System markers Glossary 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Index A Add Reference Data command 4 17 Add Sample to Project dialog box general information C 36 Add Samples dialog box Edit Menu C 37 Files tab C 40 View Menu C 39 Add Samples Tab view elements C 19 Add samples to the Project procedure C 37 advanced mode peak detection options B 17 ranges options B 14 size calling method options B 16 smoothing and baselining options B 15 algorithms allele calling B 2 B 5 binning B 2 B 5 described 1 3 peak detection B 2 B 3 quality value determination B 2 B 7 size matching B 2 size matching size callin
184. marker size range The electropherogram being viewed is scaled to the size range available for the selected marker Scale to This setting allows the user to define the horizontal range being viewed To zoom in to a specific region on the X axis of the electropherogram Click and drag with the left mouse button the desired region on the X axis OR Right click on the X axis to open the X Axis Scale menu command options Y Axis Scale Y cale Scale to maximum Dyes gt Scale to The Y Axis Scale menu command controls the vertical scale of the reference data electropherogram being viewed as follows Scale to maximum The electropherogram being viewed is scaled to its maximum height as Scale to The user can set the Y Axis scale value Using Panel Manager 4 21 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 4 22 Using Panel Manager Artisan Technology To zoom in to a specific region on the Y axis of the electropherogram Click and drag with the left mouse button the desired region on the Y axis OR Right click on the Y axis to open the Y Axis Scale menu command options Dyes Blue Dye Ctrl 1 v Show SNP Bins v Green Dye Ctrl 2 v Yellow Dye Ctrl 3 v Red Dye Ctrl 4 v Orange Dye Ctrl 5 v All Dyes Ctri 6 The Dyes menu item controls the dye colors shown when viewing SNaPshot reference data and microsatellite referenc
185. matrix to raw data allows you to generate multicomponented data Creating and Evaluating a Matrix 9 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 TAMRA ROX ome a aoe J Tr T my EMISSION 450 500 550 600 650 WAVELENGTH am 9 4 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sample Files Using The following figures show examples of data analyzed with and without A Matrix a matrix You can see that peak data from a sample file analyzed without a matrix displays the expected peak along with extra peaks in other dye colors or bleed through from other dye colors 12 50 Lane Demo Sample 4 1700 1800 1900 2000 2100 1700 1800 1900 2000 2100 2200 OEA a Sample file analyzed with a matrix Assigning a Matrix Normally the matrix is assigned to a sample file automatically upon to a Sample generation during or after a run Additionally a matrix can be manually assigned to a 377 310 sample file from within the GeneMapper software When to Assign a Before you can successfully analyze 310 377 sample files u
186. mmand 4 7 dyes 4 22 Edit Bin command 4 14 Edit Bin dialog box 4 15 Edit SNP Marker command 4 12 Full View command 4 20 Import Bins Options dialog box 4 9 Import Export Bin Set command 4 8 Import Export Panels command 4 8 Navigation pane 4 5 New Bin Setcommand 4 16 new marker 4 7 new panel 4 7 overview 4 2 Panel SNP Reference Datacommand 4 18 purpose of 2 12 4 2 rules for imported and exported files 4 32 Show Project Alleles command 4 17 toolbar 4 4 View menu 4 20 window commands 4 6 X Axis Scale menu command 4 20 4 21 scale to 4 20 Y Axis Scale 4 21 Panel Manager window 4 2 displaying 4 3 Panelnames 4 34 Panel table columns 4 34 Panel Table view 4 33 parameter values 5 7 5 9 Pass or Fail quality metrics 5 17 PCR Single stranded Conformation Polymorphism SSCP Minimum Peak Half width setting B 12 B 17 peak detection optimizing sensitivity B 23 B 25 B 27 parameters B 19 peak window size B 19 polynomial degree B 19 slope threshold B 28 peak detection algorithm advanced 5 12 basic 5 11 classic 5 13 Peak Detector tab advanced peak detection algorithm 5 12 basic peak detection algorithm 5 11 classic peak detection algorithm 5 13 Peak Quality tab allele number 5 15 Index 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com heterozygote balance 5 14 peak morphology 5 15 pull up peak 5 15 signal level 5 14 SNP 5 15 plot settings default profiles 8 2 Plot Sett
187. mple Comment field w ea LM Ej LM o LM When you add samples the panel for all samples is set to the value in the Sample Sheet comment column Project Window Software Interface C 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Add Samples Tab Elements continued Element Description Set Sample Type to 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 Sample type for all samples option button and drop down menu Read from Data When you add samples the GeneMapper collection Info field software does a best guess match comparing option button 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 Ifa match is not found then the Sample Type property for the sample is set to Sample Note The Panel and Sample Type properties accounts for close spellings and abbreviations as shown in the following table Spellings and Abbreviations Case Result A sample file field contains The sample type for the sample is set to either Control Ctrl Cri Cntrl Cont
188. mple files results and project name from the Project 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 Always enabled except when a blank Project is displayed Open Project Ctrl O Displays the Open Project dialog box If previous Project has pending changes the following alert message is displayed you want to save changes Yes No Cancel Always enabled Save Project Ctrl S Saves Project to the file named at the start of analysis Named projects are saved to the GeneMapper 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 contains data delimited text file Print Displays the Print dialog box Enabled when the tai AP The standard Print Setup dialog is ag table contains opened from the Print dialog Log Out Closes the Project window and Always enabled displays the Login window Displays a Save alert message if the Project has pending changes C 10 Project Window Software Interface Artisan Technology Group Qual
189. n Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Software Warranty Information F 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Glossary Allele Allele calling Bin Bin set Diploid polyploid Genotype GM HID Kit LMS Marker Microsatellite Panel Project SNaPshot Multiplex Analysis SNP a variant of a marker identification of alleles based on bin definitions genotyping GeneMapper software analysis a fragment size or basepair 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 usual
190. n Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com table below Window Elements The Size Match Editor window elements are described in the following 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 on a peak selects it single 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 button Allows the user to override the Sizing Quality value and automatically set the value to 1 This will cause the sizing quality to be invalidated and a check mark will display in the SQI column in the Project window Samples tab Edit View Tools Editor Commands The following Size Match Editor menus a
191. nables you to choose sample files from local or mapped networked storage devices Files pane Samples To Add pane l AddSamplestoProect OT Salse a Poed a ae a x Edit View Files GM Database Sequence Caller fer Samples To Add E _uninst3 E E E app H O Database i docs El Examples A PanelData i 0 AmpFLSTR_Panels_v3 Es gt SampleFiles H JRE 1 2 GMdistribution f y 1386 re OracleGM Ej RECYCLER B E H E F Add To List gt gt Add Samples to Project Window Elements Element Description Files pane Contains the directory structure configured for the computer running GeneMapper 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 Contains the folders and files that the system will add pans This field also uses the tree viewer interface You can use Shift click and Ctrl click as described above Add To List Adds the selected items from the files pane to the button 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 Projec
192. nalysis Methods Table Settings Plot Settings Matrices Size Standards Name Last Saved Owner Description Default 200 3 Factory Provided 2 HID Default 2002 08 20 13 0 gm Factory Provided 3 SNP Default 2002 08 20 13 0 gm Factory Provided 4 Microsatellite Default 2002 08 20 13 0 gm Factory Provided New Open Save AS Import Export Done 6 2 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Table Settings allow you to determine what columns are viewed in the Samples and Genotypes tab and to set filtering properties The Table Settings tab contains a list of Table Settings in the database This tab is used to manage the contents of the Project window by providing the following menus and interface elements Parts of the Table Settings tab Item Description Name column Table Setting name Last Saved column Date Time stamp displaying when the object was last saved Owner column User name Description column Description of the object 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 xml file Displays a dialog box for Importing objects
193. ndow size the baseline to ride too low resulting in elevated and possibly not baseline resolved peaks Size Calling GeneMapper software uses these methods to determine the molecular Method Options length of an unknown fragment Description of the size calling method parameter options Item Description 2nd Order Least Squares and 3rd Both Least Squares Methods use Order Least Squares 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 B 16 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Peak Detection Options 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 software If you leave the default Thresholds reports to the user only value of 50
194. nerated from the Instruments following instruments gt gt gt gt o ABI PRISM 310 Genetic Analyzer ABI PRISM 377 DNA Sequencer ABI Prism 3100 Avant Genetic Analyzer ABI Prism 3100 Genetic Analyzer ABI Prism 3700 DNA Analyzer Applied Biosystems 3730 DNA Analyzer Applied Biosystems 3730xl DNA Analyzer for 48 capillary instruments only Supported The GeneMapper software v3 0 is specifically designed to work with the Chemistry Kits following applications and reagents gt gt gt gt gt o ABI PRISM Linkage Mapping Set v 2 5 ABI Prism Mouse Mapping Primers v 1 0 ABI PRiSM SNaPshot Multiplex Kit ABI PRiSM SNaPshot Primer Focus Kit AmpFLSTR Kits StockMarks Animal Parentage Typing Kits ABI PRISM GeneScan Size Standards Custom Microsatellites 1 2 About GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Unique Features of Several new features and enhancements of GeneMapper software v3 0 GeneMapper_ include Software v3 0 4 Display of polyploid genotypes Remote database connection Microsoft Windows 2000 operating system support A clean installation along with upgrades for the 3100 and 3100 Avant Data Collection computers Full integration with the Applied Biosystems 3730 DNA Analyzer including automation capabilities Expanded technical portfolio i e m
195. new Bin Set Cancel 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 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 tg Delete Bin Set E x 2 This action cannot be undone Do you want to continue 4 16 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 genotypes exist 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 allowing the user to associate data with a microsatellite panel or SNaPshot kit for creating markers and bins Add Microsatellite Reference Data E x Edit Samples To Add Project Sample Analysis x Search Results Search r a Microsatellite Tutorial J Cancel For microsatellite panel reference data samples must have been analyzed using the panel for which the data serves as a reference and
196. ng and out of bounds bins are deleted 3 Delete all overlapping bins and out of bounds bins Marker Range 100 110 Marker Range 100 110 A B C D EF G If all the bins in an import are inside their marker ranges the dialog box does not open Backing up Panels and Bin Sets 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 See Export All Kits on page 4 11 4 10 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Export All Kits The Export All Kits command opens the Export All Kits dialog to allow exporting all kits and their associated panels markers and bin sets toa 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 Print The Print command opens the Print dialog box to enable printing Edit Menu The Edit menu contains the Undo command and the Clear command ig Bins View Undo The undo command allows for the Clear command to be undone Clear The Clear command deletes items selected in the navigation pane from the GeneMapper database such as Kits Deletes the kit and all bin sets associated with that kit Panels Markers Use the Bins Menu items to delete bins and bin sets Using Panel Mana
197. nment and determine if an allele should be called Bin assignment quality values are assigned to the allele and become part of the overall PQVs Questionable alleles are appropriately marked with either a Check A or Low Quality status marker See the table under Genotype View Columns on page C 7 for more information on how this information is displayed in the Genotypes view 4 40 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Analysis Methods Chapter Overview Introduction This chapter provides information on how to use the Analysis Methods feature in the GeneMapper Manager window of the ABI Prism GeneMapper Software Version 3 0 and describes how to use the features of the Analysis Method Editor In This Chapter This chapter contains the following topics Topic See Page Analysis Methods Tab 5 2 Analysis Method Editor 5 5 Using Analysis Methods 5 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Analysis Methods Tab Introducton 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 Default Analysis GeneMapper software v3 0 contains the following standard Applied Methods Biosystems profiles Dete
198. notypes 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 Show All Rows Ctrl F v Header Show Selected Rows Ctri G v Marker Range Ctri R Hide Selected Rows Ctri H v Marker Indicator Ctri K Show Allele Call Rows Ctrl V Show All Rows All available data rows are shown Show Selected Rows Only peaks that are selected in the electropherogram s are shown Hide Selected Rows All peaks that are selected in the electropherogram s are hidden Show Allele Call Rows Only peaks that have allele calls assigned are shown This is applicable for the Sizing Table only 7 20 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item 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 Headers Turned On Fe Samples Plot zax File Edit View Tools Alleles Help Mcosaiem Panes fe EE EEN ETE wee eal Mt Mue Mee Sample File Sample Name Pand os sQ 54 6 78
199. ns 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 for standard fragments that may run anomalously Peak Detection Use the Peak Detection parameter options to specify the minimum peak Parameter Options 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 Description of the 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 If you leave the default value of 50 peaks with amplitude above 50 are analyzed and display in the tabular data For each dye the GeneMapper software detects peaks above the threshold entered in the entry field Lower amplitude peaks still display in the ele
200. ns 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 hardware 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 F 2 Software Warranty Information Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Limited Product Warranty Limited Warranty Applied Biosystems warrants that for a period of ninety 90 days from Warranty Period Effective Date Warranty Claims the date the warranty period begins its ABI PRISM GeneMapper Software Version 3 0 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 di
201. nt lengths nucleotides for the nine fragments Fragment Fragment Length Length 15 62 20 80 25 110 35 120 50 Electropherogram The following screen shows an electropherogram of GeneScan 120 LIZ of GeneScan 120 run under denaturing conditions ty Size Match Editor Edit View Size Standard Size Match Quality 1 0 80 62 Te 2436 2592 2748 2904 3060 3216 3372 3528 3684 3840 3996 D 2 GeneScan Size Standards Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneScan 350 Size Standard About This Size Standard How It Is Prepared GeneScan 350 Molecular Lengths Electropherogram 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 Psil digestion of plasmid DNA followed by ligation of a TAMRA or ROX labeled 22 mer oligodeoxynucleotide to the cut ends A subsequent enzymatic digestion with BstUI yields DNA fragments containing a single TAMRA or 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 L
202. ny pending changes and closes the plot Window window Escape 7 6 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Edit Menu The Edit menu command provides basic editing for the Plot window menus described in this section Edit Item Description Undo The Undo command is enabled after a user action that adds Ctrl Z modifies clears or reorganizes It allows you to undo certain actions such as Editing an Allele Editing a Bin Resizing Markers Zooming Using Plot Windows Samples and Genotypes 7 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com View Menu The View menu contains features you can use to control zooming of electropherograms electropherogram lines dye displays on the view a number of display activities im Tools Alleles Help v Toolbar Sample Info Ctrl F1 Raw Data Ctrl F2 EPT Data Ctrl F3 Full View Ctrl Axis Scale gt Y Axis Scale Labels gt Y Bins Ctrl Shift B Peak Positions v Allele Changes Controls To Top v Off scale Peak Indicator Dyes Plots Tables Table Filter v Header v Marker Range Ctri R v Marker Indicator Ctri K vvv v arker Warg Item Description Toolbar Controls whether or not the Plot toolbar is displayed at the top of the Plot window This item
203. o Edit allele calls Genotypes Plot Window The Genotypes plot window allows you to View electropherograms on a per genotype basis View all genotype calls for a given marker Edit markers and bins View the relationship between controls and samples gt gt gt Edit allele calls Displaying Plot Display the plot window from the GeneMapper Project window Windows To open the plot window Step Action 1 Highlight any number of samples or genotypes from the project window by selecting the row number or sample file name 2 Click j Display Plots on the toolbar or Analysis gt Display Plots When you select 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 7 2 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Customizing Plot Windows 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 failed sizing quality SQ value cannot be displayed in the Plot window Only raw data can be viewed for failed samples
204. odes The Size Note All Using the Size Match Editor Match Editor window uses the mouse for editing 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 Step Action 1 Import and analyze samples in the Project window Sort failed samples to the top of the Samples tab view Select the samples whose size match you want to edit 2 3 4 Select Analysis gt Size Match Editor The selected samples display in the navigation pane at the left side of the window Selecting a sample name in this list displays its Size Standard electropherogram in the plot panel 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 10 16 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To edit a particular Size Standard peak continued Step Action 7 If you choose 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
205. of a selected panel and places it in the same kit as the original with the name Selected 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 Using Panel Manager 4 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 4 8 Using Panel Manager Import Export Panels Import Panels Lookin Paes a c a Install Standards Panels t4t a LMS HD5 2 5 Panels bet a LMS MD10 2 5 Panels a Microsatellite Tutorial Panel tt File name Panels Import Files of type all Files igi x Cancel The Import Panels command and the Export Panels command open the corresponding dialog box to allow importing exporting of kits and panels This is enabled when the Panel Manager root node is selected in the navigation pane See Formats of Panel and Bin Text Files for more information Export Panels is not available when the Panel Manager root node is 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 is enabled when a kit name is selected in the navigation pane See Formats of Panel and Bin Text Files for more information When bins are imported into a kit GeneMapper software ch
206. oftware Feature Description Ability to read genetic analysis data produced by earlier ABI PRISM genotyping applications You can import sample files directly into the GeneMapper database from either the ABI PRiSM Data Collection software or Sequence Collector database It is not necessary to use separate analysis tools such as GeneScan software and Genotyper software for peak quantitation and allele calling Note GeneMapper software has read only access to the Sequence Collector Version 3 0 database Therefore GeneMapper database results cannot be written back to the Sequence Collector Version 3 0 database Instead they are stored in individual GeneMapper projects within the GeneMapper database Ability to read and process sample fsa files containing a fifth dye Provides fourth and fifth dye support Data containing a fifth dye can be analyzed just like the other four dyes Ability to export results in a user defined format The final results from the GeneMapper software table can be exported as user defined text files Ability to import panel marker and Bin definitions Marker and bin definitions can be imported into the GeneMapper database using tab delimited text files Ability to export panel marker and bin definitions Marker and bin definitions can be exported from the GeneMapper database as tab delimited files 1 6 About GeneMapper Software Artisan Technology
207. ollowing ways Select Tools gt GeneMapper Manager gt Plot Settings Tab gt New or Open 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 Plot Window Name New Plot Setting Description Cancel 8 4 Using the Plot Settings Editor Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Sample File Sample Name Sizing Quality The Show button displays all selected rows as visible checked and the Hide button displays all selected rows as hidden unchecked 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 ample File IZ SampleName O iv E i wo oo P one Basepair anere 0n E singe Peak aot Z l aw Paak Haint Bl2lalele lele The Show button displays all selected rows as visible checked and the Hide but
208. om 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 OK Accepts the pending changes and closes the dialog box Validation the following criteria apply The edited bin must have a name unique in its marker The edited bin must have a location and offsets that do not overlap any other bins in the marker If validation fails an alert message will be displayed Cancel Closes the dialog box without accepting the pending changes Using Panel Manager 4 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Element Description New Offset Checking this box sets a left and right offset default to Default checkbox be used for subsequent creation of bins Mutant Bin Designates a bin as a mutant Bin color is red New Bin Set The New Bin Set command opens the New Bin Set dialog box allowing the user 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 SNP kits can only contain one bin set tg New Bin Set E xj i Please enter name of
209. on Guaranteed 888 88 SOURCE www artisantg com Registering When you are ready to use the GeneMapper software follow the GeneMapper instructions in the table below for registration Software To register GeneMapper software Step Action 1 Click Start gt Programs gt Applied Biosystems gt GeneMapper The first time you start the GeneMapper software the Product Registration dialog box opens og GeneMapper Eg Product Registration Your Name Organization Registration Code 2 Enter your name organization and registration code and click OK Note The first time you use the application you are asked to enter the registration code found on your registration card GeneMapper software then verifies the code 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 2 4 Using GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Logging on to and Logging out of the GeneMapper Software Introduction Logging on to the GeneMapper Software This section provides information on logging on to and logging out of the GeneMapper software You must log on to the GeneMapper software program with a user name so that the system can log information to the database and preserve the opt
210. on Slope Threshold for Peak Start and Slope Threshold for Peak End Parametets s 035 0s 0 a8 v ese ies ee a seat Lee ws B 28 Slope Threshold Examples 0 0 0 0 cece eee eee eee B 30 Size Calling Methods for Classic and Advanced Modes B 32 C Project Window Software Interface Appendix Overview 0 00 eee cence eee C 1 Overview of the Project Window 0 0 00 eee eee eee C 2 Project Window Menus 0 0 cece ee eee ee C 9 Project Window File Menu Dialog Boxes 000 C 27 Printing Tables and Electropherograms 020000005 C 34 Add Samples to Project Dialog 0 0 00 eee eee eee C 36 Project Window Edit Menu Dialog Boxes 0 0 C 46 D GeneScan Size Standards Appendix Overview nes seces cea iee eee Eie a ieke D 1 GeneScan 120 Size Standard 2 0 0 cee D 2 GeneScan 350 Size Standard 2 0 cee D 3 GeneScan 400HD Size Standard 0 0 0 0 cece eee D 4 GeneScan 500 and GeneScan 500 250 Size Standards D 6 E Sample File Conversion Appendix OVErVIEW cia kee os ye eed ee de eee ede ees E 1 Converting Macintosh Sample Files 0 0 0000 04 E 2 vi Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com F Software Warranty Information Appendix Overview roope eee cent eee Computer Configuration 0 0 0 2 cece eee eee
211. on the application when creating a new matrix for data collection Note This chapter applies only to the ABI PRISM 310 Genetic Analyzer and the ABI Prism 377 DNA Sequencer instruments In This Chapter Topics in this chapter include the following Topics See page About the Matrices Tab 9 2 About Matrices 9 3 Process of Creating a New Matrix 9 8 Choosing a Data Point Range for the Matrix Calculation 9 10 Generating a New Matrix 9 12 Assigning the Matrix to Samples 9 16 Evaluating the Matrix 9 17 Using the Matrix with Data Collection 9 18 Causes for Bad Matrices 9 19 Creating and Evaluating a Matrix 9 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About the Matrices Tab Purpose of the Matrices Tab Matrices Tab the Matrices Tab contains the list of Matrices in the database Item Description Name column Matrix name Last Saved column Date Time stamp showing when the matrix was last saved Owner column User name of the person who created the matrix of Dyes column The number of dyes selected 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 Import b
212. onowanrewss A e e P Sesana foroaren OM Matrx FS Showa Recorda Using Table Settings 6 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 g Table Setting Editor E eere fonwes afe _ SarmieNare _ srowatrecwes _ E meo orowarecoas Fi Rinname a Pet ievowatnecons l 6 6 Using Table Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The Sample and Genotypes Tab 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 Element Description Column settings These settings contain the valid columns for either Samples or Genotypes data Show column List with an editable check box for each Project column controlling whether the column is shown or hidden Column column List of column headings for the Project window Filtering column List of filtering properties for each Project window column Content column Sets what information is displayed in each Project window column as selected
213. oolbar IV Show Off scale For Sample plot only eT ela EE For Genotype plot only Marker Margin 5 bp F i Cancel 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 way these views display with the current settings on this page These icons are the same as the ones in the 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 Using the Plot Settings Editor 8 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Creating and Evaluating a Matrix Chapter Overview Introduction This chapter describes the process of creating and evaluating a matrix With the ABI Prism GeneMapper Software Version 3 0 matrices are stored in the GeneMapper database They become mtx files on your hard drive only when you export them You must export a file in order to run the 310 377 data collection You have the option of selecting four or five dyes depending
214. or unknown for linkage and SNP SNaPshot applications OL Off ladder for Human Identification HID applications blank for no allele calls Note 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 Project Window Software Interface C 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotypes View Columns continued Column Description ADO Allele Display The box is labeled with an X when the Overflow number of alleles the marker calls exceeds the number to display previously set by the user The user specifies how many alleles to display in the Table Settings parameters The default is two There are six columns for each allele to indicate name size height area mutation and comment AE Allele Edit The box displays unlabeled immediately after analysis The box is labeled with an x when the marker allele calls have been edited by the user The marker allele calls can be edited in the Plot window The use
215. ote Locations of the list and table are pointed out in the figure on the previous page Criteria selector list list of properties columns in the project window grouped by topic Field column Text field that contains the properties columns in the selected group criterion 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 Criteria column Field for specifying the text string associated with the search rules which are case sensitive Rules for Search Criteria accumulate as they are entered in different criteria 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 C 42 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GM Database Tab Elements 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 database
216. ou close the GeneMapper software using the Logout command the login window reopens after you select either Yes or No GeneMapper software closes completely if you use either the Exit command or the Close button Using GeneMapper Software 2 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Importing Data from the Analysis Sample Sheet Introduction The sample sheet output from the ABI PRISM instrument used to collect data provides important input to GeneMapper software to identify the lane number and contents of each sample to undergo analysis This section identifies the sample sheet information used by GeneMapper software so that you can correctly set up the sample sheet For detailed information on how to fill out a sample sheet correctly see the user s manual for the appropriate ABI PRISM instrument Transfer of Data is transferred from a sample sheet into GeneMapper software as Information described in the following table Sample Sheet GeneMapper 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 software Sample Info or Color Info Sample type Control Allelic Ladder Sample Negative Control Comment Panel name Comment after Panel name Use the User Defined Columns 1 3 pipebar to separate text into user defined columns Note The 373
217. p GeneMapper Project Microsatellite Project oll File Edit Analysis View Tools Help ere SA WMU MME P Table Seting SNP Defaut y 03 Samples Genotype Status Sample File Sample Name Comments Sample Type Analysis Method Panel 1 i 3fsa 2 None Sample SNaPshot Default None 2 3_C03_05fsa 3 None Sample SNaPshot Default None GS8120LIZ 3 _E03_09fsa 5 None Sample SNaPshot Default None 68350 4 6_FO3_11 fsa 6 None Sample SNaPshot Default None 65500 5 liz_C04_06 fsa liz None Sample SNaPshot Default None GSS500LIZ 6 Pe 01_100301 fsa 100301 None Sample Microsatellite Default None GS120LIZ T M 02_100405 fsa 100405 None Sample Microsatellite Default None G8120LIZ 8 Pe 03_100501 fsa 100501 None Sample Microsatellite Default None G8120LIZ 9 D 04_100302 fsa 100302 None Sample Microsatellite Default None GS 120LIZ 10 _ 05_100406fsa 100408 None Sample Microsatellite Default None 68350 1 06_100502 fsa 100502 None Sample Microsatellite Default None G6s8500 analysis Completed Lo Me j Samples View Column Descriptions Column Description Sample or row From application used to select a single entire row number 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 The
218. pen 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 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 fe iy Brings samples with errors to the top of the table sorts by overall confidence value in the Samples or Genotypes table Tooltip Low Quality to Top Ctrl B Opens the Table Setting Editor for the currently selected table Tooltip Table Setting Editor Ctrl T Opens the Find dialog box Tooltip Find Ctrl F wael Opens the Print dialog box Tooltip Print Ctrl P Using the Project Window 3 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Viewing the The following table lists how to display information in a Project window Project Window Contents To display information in a Project window To hide the
219. played when viewing Primer Peaks the raw data of your matrix standards Any time you run dye labeled samples on a gel 377 instrument or capillary 310 instrument 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 Creating and Evaluating a Matrix 9 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Generating a New Matrix Creating a To create a GeneMapper software matrix GeneMapper Software Matrix SteP_ Action 1 Select GeneMapper Manager gt Tools Then click the Matrices tab The Matrices page opens GeneMapper Manager 9 12 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To create a GeneMapper software matrix continued Step Action 2 Click the New button on the Matrices tab The Matrix Editor dialog box opens fg Matrix Editor poo poo mE ooo 00000 00000 3 Enter in a Matrix name and description if desired Creating and Evaluating a Matrix 9
220. ples and Genotypes Matrices Chapter 9 Creating and Evaluating a Matrix Size Standards Chapter 10 Using the Size Standard Settings Using GeneMapper Software 2 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Overview of the User s Manual Organization of This manual provides technical information to support the GeneMapper the User Manual software user The following chapters explain how to use the major application components Chapter 3 Using the Project Window describes how to use the Project window including the Project Manager to delete and rename Projects Chapter 4 Using Panel Manager describes how to use the Panel Manager to manage all of the chemistry kits panels markers and bin definitions required for automated allele calling Chapter 5 Using Analysis Methods describes how to use the Analysis Methods tab to create custom analysis methods for use in analyzing sample files Chapter 6 Using Table Settings describes how to use the Table Settings tab to create new profiles hide and show table columns and filter the table entries rows in Project windows Chapter 7 Using Plot Windows Samples and Genotypes describes how to view data graphically and edit allele calls Chapter 8 Using the Plot Settings Editor describes how to create a custom plot profile for viewing data Chapter 9
221. quares 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 B 38 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Software Interface Appendix Overview Introduction This appendix gives detailed descriptions of the features of the ABI Prism GeneMapper Software Version 3 0 application including details of each major application window In This Appendix This appendix contains the following topics Section See Page Overview of the Project Window C 2 Project Window Menus C 9 Project Window File Menu Dialog Boxes C 27 Printing Tables and Electropherograms C 34 Add Samples to Project Dialog C 36 Project Window Edit Menu Dialog Boxes C 46 Project Window Software Interface C 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Overview of the Project Window Menus and Dialog The Project window has a number of menus and dialog boxes Boxes associated with it The drop down menus available 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 C 2 Samples View
222. r 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 Using GeneMapper Software Note Refer to Appendix A for PQV column information C 8 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Menus Introduction The tables in this section describe the following Project Window menus Topic See Page File Menu C 9 Edit Menu C 11 Analysis Menu C 12 View Menu C 14 Tools Menu C 16 Help Menu C 26 Command The commands in the Project Window menu can be activated in two Activation and ways Special Terms Clicking the command in the menu or Using the special key combinations shown to the right of the name in the menu File Menu The commands are described in the table below Edit Analysis View Tools New Project Ctri N Open Project Ctrl 0 Save Project ChiES Save Project As Add Samples to Project Ctrl l Export Table Ctrl E Print Ctrl P Log Out Exit Alt F4 Project Window Software Interface C 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Project Window File Menu Commands Item Description Enabling New Project Ctrl N Clears sa
223. r navigation pane as follows 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 Enter in a unique kit name Select the appropriate kit type microsatellite or SNP from the drop down menu Click OK to add kit to navigation pane 4 6 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com New Panel This menu item is enabled when a kit folder is selected in the navigation pane and allows creation of a new panel folder When creating a new panel Select the New Panel name in the right hand window table and enter in the desired name Names must be unique within the database Repeat to add multiple panels to a kit Press Enter then click Apply after entering a Panel name to update the navigator pane New Marker This menu item is enabled when a panel folder is selected in the navigation pane and allows creation of a new marker When creating new markers 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 for more information Repeat to add multiple markers to a panel Press Enter then click Apply to update the navigator pane Duplicate Panel The Duplicate Panel command creates a copy
224. rameter The following are the Ranges parameter options Options Item Description Analysis Used to analyze all the data collected on the genetic drop down menu analysis instrument for each sample Full Range Enter Start and Stop data point numbers in the 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 analysis range to start after the primer peak Partial Range 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 fields drop down menu in order to specify only a limited range analyzed for All Sizes each sample Partial Sizes Software Genotyping Algorithms B 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Data Processing The Data Processing parameter options specify how the raw data is Parameter Options Processed before peak detection and size calling Description of the data processing parameter options Item Description Baseline checkbox Used to automatically adjust the baselines of all detected dye colors to the same level for a better comparison of relative signal intensity Multicomponent checkbox Note Do not che
225. ranteed 888 88 SOURCE www artisantg com Analysis Tab Elements continued Element Description If only one allele is If this option is checked single alleles will be labeled in a genotype displayed in the Genotypes table and in the Plot then duplicate the label window as two labels to indicate presumed group box homozygotes Duplicate homozygous alleles checkbox Project Window Software Interface C 23 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Users Tab The Users tab contains preferences associated with managing registered users Startup Add Samples Sequence Collector Analysis Users r GeneMapper Users UserName Created On Show mo 2002 03 28 1 cd New User H Cancel Note Once created user names cannot be deleted Users Tab Elements Element Description User Name field Displays the list of registered users User Name column Name of the user Created On column Date user was created Show column Shows the user 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 software Shift click the column label headers to sort the list by that column C 24 Project Window Software Interface Artisan Technology Group Quality
226. rd parameters Step Action 5 Click Select Sample button to select the sample and click OK The Size Standard Editor opens kp Size Standard Editor 2334 3112 3890 4668 5446 6224 7002 7780 8558 Using the Size Standard Settings 10 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using the Size The Size Standard Editor window in Basic or Advanced mode allows Standard Editor in you to define the peaks for each size standard dye you use Basic or Advanced Mode La Size Standard Editor To use the size standard editor in basic or advanced mode Step Action 1 Choose a Size Standard Dye color from the drop down list 2 Enter a description for the size standard you are creating 10 8 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To use the size standard editor in basic or advanced mode continued Step Action 3 Enter the base pair values to be defined in your size standard in the table provided Note Use the Edit menu to undo certain actions or Add and Remove Rows as desired Edit Add New Row Ctrl A Remove Row Ctrl R Click OK to save and close the Size Standard Editor or click Cancel to close without saving Using the Size Standard Settings 10 9 Artisan Technology Group Quality Instrumenta
227. re are other analysis options which ignore the status field The Status field next to each sample contains the Lan symbol prior to analysis This field is empty after analysis if analysis was successful 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 Editable free text Project Window Software Interface C 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Samples View Column Descriptions continued Column Description Sample Type Editable via Sample Type drop down list box valid types are Sample Control Allelic Ladder and Primer Focus Negative Control Analysis Method Editable via a drop down list box The list is editable via the Analysis Method Editor in the Analysis Methods tab in the GeneMapper Manager window Panel Editable via Panel window Panel window contents come from Panel Manager Size Standard Editable via a drop down list box The list is editable via the Size Standard Editor in the Size Standards tab in the GeneMapper Manager window Matrix Editable via a drop down list box The list is editable via the Matrix Editor in the Matrices tab in the GeneMapper Manager For the ABI PRIS
228. re described in this section Using the Size Standard Settings 10 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Edit Menu The Edit menu is used to delete or change size labels iia View Tools Mag Pad size lapel Delete Size Label Delete Change Size Label Delete All Size Labels Ctrl Delete Item 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 selected Enabled when an Label peak unlabeled peak is selected Delete Size Deletes the size label of the Enabled when a Label selected peak labeled 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 labels for Always enabled Size Labels the selected size standard sample Ctrl Delete 10 14 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com View Menu The View menu is used to change the Size Match Editor window Hena Tools zoom gt Y Axis Scale gt v Show Navigator Ctrl Shift N Item Description Enabling Zoom Submenu used to scale the plot to Always enabled the following
229. ree 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 a Degree Value Value Increase sensitivity use Higher Lower Decrease sensitivity use Lower Higher Software Genotyping Algorithms B 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Guidelines for To detect well isolated base line resolved peaks use polynomial Using These degree values of 2 or 3 For finer control use a degree value of 4 or Parameters 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 h
230. roject quickly then click the Open button to open the selected project or double click the project name gr Open Project coe 12 Microsatellite Project 2002 08 2 om Casework Project 2002 06 05 14 59 55 a Bo Casework Project 06 2002 06 05 15 22 29 BT E HID Casework 0626 2002 08 01 15 33 47 oe __ _ ee ee 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 C 28 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The following table describes the Open Project dialog box window elements Open Project Window Elements Element Description Search field Typing in this field selects the first item in the project list whose name matches the characters Project table Lists all the projects in the database Project Name of the project Last Saved Date and time stamp of when the project was last modified Owner Name of the user who created the project 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
231. rol or a similar variation C 20 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sequence Collector Tab The Sequence Collector tab enables access to a single Sequence Collector Startup Add Samples Sequence Collector analysis Users Database Connection Test Test Result NotTested User Name Password Doo o O C Save Database Schema Owner H Cancel Sequence Collector Tab Elements Element Description Test Result The results returned are Pass or Fail These results are displayed by clicking the Test Connection button If the Test Connection button has not been pressed the Test Result will display Not Tested User Name Registered user name to access Sequence Collector Password Password for the user name to access Sequence Collector data entry into this field displays as bullets or asterisks Save Save the Password when set the user does not have to check box enter a password to get access to Sequence Collector Database Name of the Sequence Collector Sequence Collector string refer to Sequence Collector documentation for more information Schema Owner The name of the owner who has access to desired sample collections Test Connection Tests the connection to the database specified includes button appropriate error checking per Sequence Co
232. rom the original pixel position When you replace the CCD camera ABI PRISM 310 Genetic Analyzer and ABI Prism 377 DNA Sequencer When you see an increase in pull up or pull down 9 6 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Considerations The following table lists some of the considerations before making a Before Making a Matrix Where to Store Exported Matrix Files matrix Consideration Comment How much dye matrix standard to load With the ABI PRISM 377 DNA Sequencer loading more than 3 uL will overload most wells and may produce too much signal Any amount that results in a signal over 4 000 RFUs is too strong Which lanes to load with the dye matrix standards For gel electrophoresis load the matrix standards with an empty lane between each sample to avoid contamination of the individual dyes by residual material leaking adjacent samples Which gel data will be used for matrix creation After generating a gel image for ABI PRISM 377 instrument check that the tracking of the gel file is adequate Store exported matrix files mtx intended for use by Data Collection software in 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 analys
233. rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Process Quality Values Appendix Overview Introduction 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 ABI Prism GeneMapper Software Version 3 0 to troubleshoot the data analysis process In This Appendix This appendix contains the following topics Topic See Page Overview of Process Component Based Quality Values A 2 PQV Using Process Component Based Quality Values A 3 Process Quality Values A 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Overview of Process Component Based Quality Values PQV What are Process Process Quality Values PQVs values reported by data analysis aid in Quality Values 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 Atthe 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 Onthe Samples page and Genotypes page the columns presenting PQV result data other than SQ and GQ results display the following samples after analysis Pas
234. rtisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Causes for Bad Matrices Ifan Error There are two possible causes for the error messages shown in the Message Diplays following table If Take this action you designated the wrong files reassign the matrices Refer to Assigning the Matrix to Samples on page 9 16 the signal is too weak to make a matrix rerun the matrix standards Two Causes of Bad The following table lists two common causes of bad matrices Matrices Problem Cause What to do Artifact peaks of different colors under the true peaks Refer to the figure below Loading too much dye when running matrix standards resulting in dye bleed through Complete another run and recreate the matrix 4B A11 Test 24 2 OW 4 att test 24 27 ve 4G Al1 Test 24 2 7 4R Al1 Test 24 2 7 Noisy baseline If the matrix subtracts too much of a particular color from the sample then the baseline may become too elevated resulting in false peaks Refer to the figure below Complete another matrix run and make sure you do not have any off scale data Or choose a different set of fragments Creating and Evaluating a Matrix 9 19 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www
235. rtisantg com Bins Definitions The examples below show SNP bin definitions in the format used by for SNaPshot GeneMapper software v3 0 in a tab delimited text file txt 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 j SNPdemo_Bins txt Notepad lol x File Edit Format Help SNPdemo SNPdemo 6plexdemo 20mer 26 1 26 1 Green 28mer 29 9 29 0 Blue 29 9 Green 36mer 40 2 40 1 Blue 44mer 48 8 48 8 Red 52mer 5S2 54 6 yellow 55 2 Red 60mer 63 0 63 0 Columns 1 bin name 2 bin size range min 3 bin size range max 4 bin color 4 30 Using Panel Manager Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Ed Microsoft Excel Book3 Ee x File Edit View Insert Format Tools Data Window Help 20mer 24 6 26 1 24 6 26 1 Green 28mer 27 3 29 9 27 3 29 1 Blue 28 5 29 9 Green 36mer 38 4 40 2 38 4 40 2 Blue 44mer 46 8 48 8 46 8 48 8 Red 52mer 53 1 55 2 53 1 54 6 Yellow 54 55 2 Red 6O0mer 61 4 63 61 4 63 Yellow Columns 1 bin name 2 bin size range min 3 bin size range max 4 bin color Using Panel Manager 4 31 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Import and Export The Panel Manager imports and exports two text file data formats Files panel d
236. 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 sla 11eP2 11 HE Best Fit 2nd Order Curve AQ 9 771623E 01 Al 1 220838E 01 A2 4 856885E 06 R 2 1 000 Size Calling Curve Global Southern Method T T T T T T T T 1000 1500 2000 2500 3000 3500 4000 4500 Data Point AES A E Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper software v3 0 Software Genotyping Algorithms B 37 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 di 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 How This Method Works All points in the standard are weighted equally and the curve is 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 s
237. s For microsatellite data Step Action 1 Select a marker in the Panel Manager navigation 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 Edit the bin information in the Add Bin dialog box that opens For SNaPshot data Step Action 1 Select a SNaPshot 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 Using Panel Manager 4 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Delete Bin The Delete Bin command deletes a marker bin as follows For microsatellite data Step Action 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 on the bin and select the Delete Bin option For SNaPshot data Step Action 1 Select a SNaPshot 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 on the bin and select the Delete Bin option Edit Bin The Edit Bin command edits
238. s 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 an acceptable response 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 the user for editing A 2 Process Quality Values Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Using Process Component Based Quality Values The PQV System Quality values reported by the GeneMapper 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 Software v3 0 on page 1 3 The color coded sortable PQV values on the Samples and Genotypes pages allow the user 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 The SFNF MNF SNF and OS parameters report results as Pass F or Che
239. s present at a position within 0 5 basepairs from a bin and no peak is present inside that bin The benefit is to capture peaks that fall outside of bin boundaries due to experimental variations This flag is used for SNP only DP Double Peak A flag Pass Ii or Check A is automatically displayed when two peaks of the same color in the same bin have a ratio gt 0 5 minor peak height major peak height This flag is used for SNP only Not editable Process Quality Values A 11 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description GQ Genotype Quality A Low Quality or red octagon is shown when the analysis has failed for that sample Pass I Check A 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 AG AG See the following page for information about the mathematical formula used to calculate the Genotype Quality from a sample represented by the image above A 12 Process Quality Values Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description GQ continued Genotype Quality Mathematical Formula The m
240. scovered 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 i provide any software corrections or bug fixes of the identified failure if and when they become commercially available to buyer free of charge or ii notify buyer that Applied Biosystems will accept a return of the software from the buyer and upon such 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 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 Any applicable warranty period under these sections begins on the 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
241. sers 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 Startup Tab The Startup tab contains preferences associated with the startup of the software Project Window Software Interface C 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Startup Tab Elements 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 Add Samples Tab The Add Samples tab contains preferences associated with the adding of samples x Startup Add Samples Sequence Collector Analysis Users When adding new samples au omatically Set Analysis Method to Cc Defauit z for all samples Read frem the Sample Set Site Standard tn s esso z for all semples Read trom the Sample Set 310 377 Matrix to eaves 7j for all semples Read from the Sample Set Panel to for all samples Read frem Data collection CommentParel field Set Sample Type to Sample z for all samples Read from Data collection Info fizld i Darvel C 18 Project Window Software Interface Artisan Te
242. sing the Matrix GeneMapper software you must make a new matrix or assign an existing one to a set of sample files Creating and Evaluating a Matrix 9 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Limitations to The matrix is instrument specific You cannot apply a matrix you made Matrices 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 PRISM 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 When to Create a Create a new matrix in the following conditions New Matrix For each dye set D F G5 E5 When you change the dye set you use to label sample fragments for example if you are using the fifth dye When you use gel materials or buffers with pH values that differ greatly from the pH value of the gel material or buffer on which the existing matrices were generated When you see multiple unexpected peaks of different colors under an expected peak When you recalibrate your CCD camera ABI PRISM 310 Genetic Analyzer and ABI PRISM 377 DNA Sequencer and the change is greater than 3 pixels f
243. sults Set up table format for export using Table Settings Editor Export using Export Table command 2 2 Using GeneMapper Software Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Installing and Registering the GeneMapper Software No Prior Oracle An important requirement for the GeneMapper software installation is Installation that no previous Oracle installation be in place at the time of installation except for 3100 Data Collection Software DCS the 3100 Avant DCS and the 3730 DCS CD ROM IMPORTANT Do not work off of the CD ROM Install the software to the Contents Selected drive GeneMapper software comes on a CD ROM and includes Microsatellite Human Identification HID marker bin definitions and example data and SNP genotyping tutorial data AppleScript sample file conversion utilities Electronic PDF version of the user s manual Size standard folder which contains various size standards gt gt gt Panel folder which contains Linkage Mapping and AmpF STR panels and bins Installing To install GeneMapper Software version 3 0 please use the instructions GeneMapper in the ABI PRISM GeneMapper 3 0 Software Installation Instructions Software P N 4338561 found in the kit contents or use the GeneMapper 3 0 Installation txt file on the CD ROM Using GeneMapper Software 2 3 Artisan Technology Group Quality Instrumentati
244. t Saved column Date Time stamp showing when the plot setting was last saved Owner column User name of the person who created the plot setting Description column A description of the plot setting New button Opens the Plot Settings editor Always enabled Open button Opens the editor for a selected project Enabled when a single plot setting is selected Save As button Displays the Save As dialog box Enabled when one or more settings are selected Import button xml file extension 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 Using the Plot Settings Editor 8 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 The items displayed in the Sample headers The items displayed in the Genotype headers The columns displayed in the Sizing table How labels are shown gt gt gt Display settings The Plot Settings Editor can be accessed in the f
245. t dialog box C 40 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GM Database Tab The GM GeneMapper Database tab enables you to choose sample files that have previously been added into a GeneMapper project This database uses a search interface to extract samples that match your criteria _Criteria Selector list Samples To Add pane _ __ Search Criteria Table amp Add Samples to Project ix Edit View Files GM Database Sequence Collector Samples To Add arch Criteria Condition starts with Project Create Date is Criteria Sample Analysis Genotypes zl Project Modification Date js Search Results Search Add To List gt gt GESni Add Hane reel za MERS ancei 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 Project Window Software Interface C 41 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GM Database Search Rules Criteria and Rules Description Search criteria This list and table contains the search criteria from which to extract samples from the GeneMapper database N
246. tch the kit panel chosen for analysis 5 8 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Marker Repeat Type for Microsatellites 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 in the Alleles page causes the algorithm to use your defined stutter ratio and not the ones defined in the Stutter ratio box Different values can be used for different markers The Cutoff value ignores all peaks less than the cut off ratio of the largest peak in the allele size range The PlusA 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 The two columns labeled Trinucleotide and Tetranucleotide allow entry of all related parameters directly into fields so that everything can be seen at once The following table provides information about the ranges of possible values Trinucleotide Tetranucleotide Parameter Min Max Default Default Cut off value 0 0 1
247. th 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 is labeled Consequently even if the two strands migrate at different rates under denaturing conditions you will not need to worry about peak splitting The following figure shows the peak patterns of GeneScan 400HD fragments run under denaturing conditions Fragments were run using the 3700 POP 6 polymer at 60 C D 4 GeneScan Size Standards Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Electropherogram The following screen shows an electropherogram of GeneScan 400HD of GeneScan 400HD Match Editor Size Match Quality 1 0 34 30 30 oa 20 280200900 100120 aso 160 18019020 720 ia 17007 50 60 100 500 100 t t 2836 3094 3352 3610 3888 M2 484 dBA 4900 158 SM6 S674 5982 GIW 64B 6706 6A 7222 740 GeneScan Size Standards D 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com GeneScan 500 and GeneScan 500 250 Size Standards About These Size The GeneScan 500 the GeneScan 500 LIZ the GeneScan Standards 500 250 and the GeneScan 500 250 LIZ Size Standards are useful for sizing fragments betwe
248. the software is delivered to the buyer Warranty claims must be made within the applicable warranty period Software Warranty Information F 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 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 B
249. the SQ to 1 0 Click Apply if you are going to edit a second size standard peak or click OK if you are finished applying your changes to the edited peak Using the Size Standard Settings 10 17 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 gy Size Match Editor k x Edit View Tools Size Matches Size Calling Curve Best Fit Second Order Curve 504 AO 45 765726443467 1 Al 0 057449780294601956 A2 9 55835197463482468 R 2 0 9999186353 Size Calling Curve Local Southem Method 600 700 800 300 4000 4100 4200 14300 4400 14500 4600 4700 1800 1300 aj ee Red Curve Ifthe size calling curve is a red curve it is based on the sizecalling 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 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
250. the appropriate peaks of the internal size standard and uses this information with the selected size calling method to size all unknown fragments Applied Biosystems provides several fluorescently labeled size standards which are described in Appendix D If necessary you can also define your own size standard Advantages of Running an internal size standard results in precise molecular length Using a Size determination because the internal size standard and the unknown Standard fragments undergo exactly the same electrophoretic forces GeneMapper software can then compensate for band shift artifacts caused by variations in the gel and in the sample from lane to lane or injection to injection 10 2 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Size Standards Tab Purpose of the Size The Size Standards tab in the GeneMapper Manager window contains Standards Tab a table of size standards stored in the database a GeneMapper Manager xi Projects Analysis Methods Table Settings Plot Settings Matrices Size Standards Name Last Saved Owner Type Description 377_G5_HID_GS500 20 asic anced Factory Provided Done Item Description Name column Size Standard name Last Saved column Date Time stamp showing when the size standard was last saved Owner column User name of the person who created the size st
251. the label box Labels are determined by the GeneMapper software v3 0 AE Comment Drop down menu item in the label box Shows the last allele edit comment Show data type Check box Label prefixes such as prefixes al for allele call will be displayed Show type of edit Check box If an allele is manually edited the label will display changed or deleted Invert mutant labels Check box If a bin is labeled as a mutant bin in the Panel Manager then the label color will be inverted to distinguish it from normal bins Label Color Drop down menu Controls the color of the labels From the drop down menu select one of the following Black amp White Dye Color All text amp border in color Dye Color Border text in black border in color 8 8 Using the Plot Settings Editor Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Display Settings The Display Settings tab controls how each plot setting will Tab automatically open when a customer chooses that setting Plot Settings Editor x General Sample Header Genotype Header Sizing Table Labels Display Settings When Opening The Plot Window C Use the display settings last used for this plot Use these display settings o For both Sample and Genotype plots Panes 1 F fea fie SETE Axis Basepairs y Axis Scale individually gt I T
252. tings icon or Tools gt Table Setting Editor 2 Select the Table Settings tab 3 Select a table setting name and click Open The selected setting opens 4 Perform edits in the table setting Note See The Sample and Genotypes Tab Elements on page 6 7 5 Click OK when you have completed editing The changes you have made are saved to the Table Setting Deleting a Table To delete a Table Setting Setting Step Action 1 Select the Table Settings tab in the GeneMapper Manager window 2 Select a table setting name and click Delete A warning alert is shown Click OK to remove the setting Using Table Settings 6 9 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Filtering Controls The 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 tg Table Setting Editor x General Samples Genotypes Filtering Content Show All Records N A Show All Records Show All Records Show All Records Show All Records Show All Records N A Show All Records Show All Records Show All Records Samples Table Settings Show Column 1 Status 2 Sample File 3 Sample Name 410 Sample ID 5 iV Comments 6 iV Sample Type 7 Analysis Method 8 M Panel 9 M Size Standar
253. tion Guaranteed 888 88 SOURCE www artisantg com Using the Size The Size Standard Editor in Classic mode shows an electropherogram Standard Editor in and a table of peaks for the dye color and sample selected The Classic Mode 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 Size Standard Editor po 1556 2334 3112 3890 4668 5446 6224 7002 7780 8558 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 10 10 Using the Size Standard Settings Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com To use the size standard editor in classic mode Step Action 1 Enter in a name and description for the size standard 2 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 electropherogram by click drag on the X Axis numbers Type the peak size in the corresponding column Note Leave a zero in the Size column
254. tisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Add Bin Allows the user to add a marker bin as follows Ctrl B a Select the Binning mode b Click the Marker Range for the bin A line and a cross hair opens on the electropherogram c Select the Add Bin menu item or right click to open the Add Bin option d 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 the user to add bin information Add Bin to D651574 x Name Bin Location 58 38 Left offset 0 40 I New offset default Right offset io 40 I New offset default I Mutant Bin OK Cancel Edit Bin Allows the user to edit a marker bin as follows Ctri U a Select Binning mode b Select the bin to be edited c Select the Edit Bin menu item or right click the selected bin to open the bin edit options and select Edit Bin The Edit Bin dialog box opens allowing the user 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 a Select Binning mode b Select the bin to be moved c Click and drag the center of the bin to a new location Using Plot Windows Samples and Genotypes 7 29 Artisan Technology Group Quality Instr
255. ton displays all selected rows as hidden unchecked Using the Plot Settings Editor 8 5 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Sizing Table Tab The Sizing Table tab controls which columns are displayed in the sizing table as shown in the Samples Plot view Plot Settings Editor yelSample Peak ample File Name DyerSample Peak Sample File Name Marker Alee o Size Height Area Wicd fil clad al Dd ea ata Point The Show button displays all selected rows as visible checked and the Hide button displays all selected rows as hidden unchecked 8 6 Using the Plot Settings Editor Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Allele Call Dye Color Border ha Times NewRoman fio E Use the drop down menus to display the labels you want to use Plot Settings Editor Allele Call gt Dye Color Border ha Using the Plot Settings Editor 8 7 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Description of items displayed on the Labels tab Name Type Description Allele Call Area Height Data point and Size labels Drop down menu items in
256. tor 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 Examples Table Printout ABI PRISM GeneMapper 3 0 TM SNP Samples Aug 20 2002 11 43 25 AM Status Sample File Sample Name Comments Sample Type 1 SNP5DyeC132 SNP5Dye None Sample 2 SNP5DyeC132 SNP5Dye None Sample 3 SNP5DyeC132 SNP5Dye None Sample 4 SNP5DyeC132 SNP5Dye None Sample 5 SNPS5DyeC132 SNP5Dye None Sample 6 SNP5DyeC1 32 SNP5Dye None Sample 7 SNP5DyeC132 SNP5Dye None Sample 8 SNP5DyeC132_ SNP5Dye None Sample 9 SNP5DyeC132 SNP5Dye None Sample 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 C 34 Project Window Software Interface Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Plot Printout ABI PRISM GeneMapper 3 0 TM Mixed Tue Aug 20 13 53 57 PDT 2002 Sarre aa a Ras RS T Sample File Sample Name Panel os sa CS name eee ee mE A Ma 0_C06 fsa Fe S605 Paneli D19255 E E E C E C L E L E INA A NA jog 104 106 108 110 112 114 116 E a a A S S a prowa Jow Setos Pana D1s2667 a E L O
257. tric columns are set to display color symbols by default However the SQ column in the Samples view and the GQ column in the Genotypes view can be set to display the numerical equivalent on which the color symbols are based for these columns This setting is made in the Analysis tab in the Options window by clicking the Numbers option button for the Quality Metrics Display setting 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 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 fails and any result from 0 26 to 0 74 will be marked for you to check See Process Component Based Quality Values in Appendix A 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 B 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 Using Analysis Methods 5 17 Artisan Technology Group Qu
258. trumentation Guaranteed 888 88 SOURCE www artisantg com Project Window Edit Menu Commands continued Item Description Enabling Fill Down Fills the selected column with the Enabled when a contents of the first cell This column is selected or Ctrl D command is applicable only to multiple cells are Analysis Method Panel Size selected Standard and Matrix columns Find Displays the Find dialog box Enabled when the table Ctrl F contains data Sort Displays the Sort dialog box Enabled when the table Ctrl G 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 em view Tools Help Display Plots Ctri L Size Match Editor Analyze Ctri R Analyze Selected Samples Analyze All Ctrl Shift R Analyze Markers Analysis Method Editor Size Standard Editor Bring Errors to Top Ctrl B Bring Non concordant Samples to Top Project Window Analysis menu commands Item Description Enabling Display Plot Opens the Plot window Enabled when one or Ctrl L more samples or records are selected Size Match Opens the Size Match Editor Enabled only when Editor window sized samples are present in the Project C 12
259. umentation Guaranteed 888 88 SOURCE www artisantg com Item Description Delete Bin Allows the user to delete a marker bin as follows Delete a Select Binning mode b Select the bin to be deleted c 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 Step Action 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 online help and to the About GeneMapper software window GeneMapperHelp F1 About GeneMapper Item Description GeneMapper Help Opens the GeneMapper User Manual PDF file using F1 Adobe Acrobat Reader About Opens the About GeneMapper window to indicate GeneMapper registration information and software version number 7 30 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Special Feature Invalidate Genotype Quality This feature allows the user 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 box will not
260. unName Panel Marker Dye allelet allele2 Sizet _ Size2 _ Heighti Height2 ADO AEH 0S SHP OBA SPA sP _inkage Panei01 1D182797 B 102 fi30 10232 129 99 1373 0 2950 O O CE EL command from 2 inkage Paneio1 1D182800 B 211 215 21105 21496 10720 7640 O 0 3_inkage Panelo1 1 D15234 B 276 282 275 02 280 88 1202 0 5450 O O the View menu 1 inkage Paneio1 1 D15249 B i79 179 37 183 31 7300 5470 O O 5 i 5 inkage Panel1 1 D15450 B 336 336 16 1577 0 io Oo or Ctrl Shift 2 6 _inkage Paneioi n oisies o 322 332 32228 33207 4100 2790 O O m m m i im 7 inkage Panei01 1018255 6 a5 agaa 2146 0 CO mmm En Note Samples must be analyzed to see genotypes To see the Click the And GeneMapper software displays the Samples view Samples tab Project in the Samples view again g GeneMapper Project Untitled Bees asn BS we Am SG Project Linkage Samples Genotypes Table Profile Applied Biosystems Defe Sample ID Comments Sample Type gt Status Sample File Sample Name 01_100301 fsa 100301 02_100405 fsa 100405 None Sample Sample Sample Sample 03_100501 fsa 100501 04_100302 fsa 100302 05_100406 fsa 100406 Sample 06_100502 fsa 100502 Sample 4 naling i A 3 14 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com
261. ur interactive website at www instraview com information on price quotations drivers technical LEASING MONTHLY specifications manuals and documentation RENTALS ITAR CERTIFIED D a gaa tia Contact us 888 88 SOURCE sales artisantg com www artisantg com
262. uses the known standard fragments and the associated data points to produce a sizing curve based on Multiple Linear Regression Advantages In the figures on page B 33 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 is strictly length dependent This method automatically compensates for fragments that run anomalously GeneMapper software v3 0 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 B 32 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 11eP2 11 Best Fit 2nd Order Curve aso AO 9 771623E 01 T Al 1 220838E 01 A2 4 856885E 06 200 R 2 1 000 150 100 so Size Calling Curve 2nd Order Least Squares T T T T T T T T 1000 1500 2000 2500 3000 3500 4000 4500 Data Point 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 software v3 0 11 P2 11 Best Fit 3rd Order Curve aso AO 1 26
263. ust 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 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 Basically the baseline value at each data point is the lowest electropherogram value in a window whose width Beta is set using the Baselining option and centered at each data point i Software Genotyping Algorithms B 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com More accurately the baseline computed in this manner is intermediate The real baseline value at each data point i is the highest intermediate value again in a window whose width Beta is set using the Baselining options and centered at each data point i GeneMapper 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 a large baseline wi
264. utation screening Descriptions of Benefits GeneMapper software using ABI instruments Linkage Mapping Set LMS AmpF STR HID markers or SNaPshot Multiplex provides capabilities that go beyond the ABI PRISM GeneScan and Genotyper software products It provides a genotyping capability that sets new standards for high throughput and automated genotyping with the following benefits Genotyping benefits of the GeneMapper software Benefit Description High accuracy for size and allele calling Sophisticated algorithms such as Multiple Allele Peak Determination MAPDA Automatic Bin Builder ABB and Automatic Bin Assignment Algorithms ABAA assure highly accurate allele calls High throughput In a recommended computing platform GeneMapper software processes over 50 000 genotypes in 1 hour This is sufficient capacity to analyze all the sample files accumulated by an Applied Biosystems 3730 DNA Analyzer run overnight at peak capacity in 1 to 2 hours About GeneMapper Software 1 3 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotyping benefits of the GeneMapper software continued Benefit Description Fully automated operation Process Component Based Quality Values PQV monitor major components of the size and allele calling process informing a user of the source of problems anywhere along the data analysis
265. utomatic Bin Builder ABB Builder and and Automatic Bin Assignment Algorithm ABAA Automatic Bin Assignment Combining Unique Features Algorithm The ABB process is used along with the outcomes of the Multiple Allele Peak Determination Algorithms MAPDA and in conjunction with the ABAA to ensure that the called alleles are of the highest accuracy The key benefit of combining these algorithms with the ABB is that since only a small number of questionable alleles result from the integrated process manual reexamination of the allele information is significantly minimized Automatic Bin Builder The ABB is the first step in accurate allele assignment The process of creating bins starts with a collection of Sample files Bins are created by the ABB based on the chosen panel information and successive allele calls from the Sample file collection As each Sample file in the collection is processed the bin definitions are refined to reflect the actual data The bin centers and widths continue to be refined until all Sample files in the collection are processed and GeneMapper software is ready for its final step of allele calling Automatic Bin Assignment Automatic Bin Assignment Algorithms Once bins are completely defined by the ABB allele peaks are accurately and automatically assigned to their corresponding bins to complete the allele calls using the ABAA The allele assignment algorithms calculate the certainty of each allele peak assig
266. utton 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 Done button Closes the GeneMapper Manager 9 2 Creating and Evaluating a Matrix Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com About Matrices Introduction Matrix Definition Multicomponent Definition There are two dye labeling chemistries currently available to prepare nucleic acid samples to use the GeneMapper software on ABI PRISM instruments 5 end labeling Dye Sets D F G5 3 end labeling Dye Set E5 Each chemistry has a set of dye labels that fluoresce at different wavelengths when excited by a laser During data collection on the The wavelengths are separated ABI Prism 310 Genetic Analyzer by a spectrograph into a known ABI Prism 377 DNA Sequencer spectral pattern across a detection 377XL or 96 lane upgrade system with the sequencer instrument A Matrix is a mathematical formula that corrects for spectral overlap of fluorescent emission spectra data collected from ABI PRISM instruments This process of eliminating the bleed through caused by spectral overlaps is called multicomponenting Applying a
267. v 3 g hog o Si Hh 1 I I I Me AIL FE A ipa k JN J Nowe d a ad hen erin k VR tae wend MA P V Aa phe J wees Wes f k Dorv fates PES EER i Wann A WY p SE cai paiaina aAa aae I I l i dl LI I ler td l 0 400 800 1200 1600 scan line B 4 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Binning Algorithm Allele Calling 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 the flexibility bins may also be edited manually Note When the user creates 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 Final allele calls are based on a consensus between several different 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 Example Output of Different Allele Calling Algorithms The following is an example of three different allele calling algorithms for 16 samples User annotations ar
268. value Full View Ctrl Y Axis Scale Scale to maximum Y default Always enabled Scale to user defined Show Navigator Switches the navigator pane Always enabled Ctrl Shift N Removes the pane with the first use and restores it on the second use Tools Menu Tools Auto Adjust Sizes Check Sizing Quality 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 Using the Size Standard Settings 10 15 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Toolbar The Size Match Editor toolbar contains the following icons Icon Description Enabling Lb Autoadjusts size standard peaks Adjusts the size of other peaks automatically to the right of the selected peak Tooltip Auto Adjust Sizes Enabled after a peak is changed g Checks the quality of the current size matching Checks the quality of the revised peak positions and updates Enabled after a peak label is changed the Size Match Quality indicator Tooltip Check Sizing Quality x Deletes all size labels Always enabled Tooltip Delete All Size Labels a Only for Basic and Advanced modes b Only for Basic and Advanced m
269. value of zero will not affect the sizing accuracy or precision for an asymmetrical peak B 28 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 Slope Threshold for Peak Start value from zero to a positive number start point of a peak closer to its apex end point of a peak closer to its apex Slope Threshold for Peak 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 Software Genotyping Algorithms B 29 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Slope Threshold Examples Initial The initial analysis with a value of O for both the Slope Threshold for Electropherogram Peak Start and the Slope Threshold for Peak End value produced an asymmetrical peak with a noticeable tail on the right side GEJ 2w aw znw zwo z0 amo 2180 2190 2200 2210 2220 220 2240 2250 220 2770 2280 290 20 RO 20 Electropherogram showing an asymmetrical peak B 30 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisant
270. w labels are viewed beneath an electropherogram Labels No Labels Y Bins Ctrl Shift B Horizontal Labels Peak Positions Vertical Labels No Labels Labels are off sil File Edit View Tools Alleles Help crosan JE Panes fr EEEE Bi Mew Seal ia Fuel Tne E Sample File Sample Name Pand ressor seo pma or m E 0 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 Samples Plot Ee iol x File Edit View Tools Alleles Help ENP Defaut Panes 2 z EE SEH Bh MaA Bae mA fire Mae Sample File Sample Name Panel os sa 5 E0 O9fsa 5 plex sanp N E ane 19 2 23 oe 29 31 33 35 37 4 B 45 eae 31 53 55 57 59 61 63 65 67 Ea Ede Gilmer aver ee Eza Eher 2504 2146 1728 aa 1073 715 357 0 jal A fal G Al jal G ial T lal CiT al c Lal SSS F 7 12 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Labels Vertical Labels Labels are positioned vertically across continued the screen to prevent overlap This option is recommended for printing inixi File Edit View Tools Alleles Help f
271. wcrosateme E Panes EEEE GG wee Seal A he Mefe Sample File Sample Name os sQ 01_100301 fsa rocsex seo pna o1 E 10 bisz7s7 men prszxoa 8 Mo 39 407 415 423 434 439 447 455 463 471 479 187 495 203 211 219 227 235 most 1 1005 304 804 703 603 502 402 301 201 100 I o 124 mA man fa 136 fa 163 ma 4 E Note Labels are enabled in Separate Dyes mode only for Microsatellite samples and are enabled in Combine Dyes mode only for SNaPshot samples Using Plot Windows Samples and Genotypes 7 13 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Bins Controls whether or not transparent bins are shown or Ctrl shift B hidden on an electropherogram In the Binning mode bins are automatically displayed and the Bins menu item is checked Separate Dyes mode lolx File Edit View Tools Alleles Help E Pwerosatenie E Panes r meme Bh we ala MA ufe Maaf Sample Tile Sample Nane Pand os sQ Eee 100301 Se 01_Pand 01 C h 0 T Deseas bisa rs200 J s2 400 4108 116 1424 432 440 448 156 464 172 180 188 19 204 212 220 228 a 122126 fal 179183 fa 213 217 SSSI E Note Bins are enabled in Separate Dyes mode only
272. www artisantg com Using the Project Window Chapter Overview Introduction This chapter provides a general overview of the Project window the central window of the ABI Prism GeneMapper Software Version 3 0 interface In This Chapter This chapter contains the following topics Topic See Page Central GeneMapper Software Window 3 2 Details of the GeneMapper Project Window 3 5 Exported Project Tables 3 20 Projects Page 3 23 Using the Project Window 3 1 Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Central GeneMapper Software Window Importance of the As shown by the figure below the Project window is the most important Project Window element in the GeneMapper software user interface The GeneMapper software is organized around the concept of the project to support its purpose of automatically calling alleles or performing genotyping Splash screen Registration screen Project Manager Analysis Methods Add Samples to Project dialog box Table Settings o Project Open and Plot Settings Save dialog boxes i Print dialog box Mat Manager Manager Samples and Genotypes Size Standard 9 p yp Export Table Editor dialog box Panel L Import Export Manae Plot window Options dialog box dialog boxes eevee Sort and Find Editor dialog boxes
273. y GeneMapper software A split peak is defined as overlapping peaks that are lt 25 basepairs apart the horizontal distance from two peak apexes Not editable OS Offscale A flag Pass Ii or Check A is automatically displayed when there are offscale peaks present within the marker size range This flag is used for all applications Not editable A 10 Process Quality Values Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com The PQV flags on the Genotypes page continued Column Description cc Control Concordance A flag Pass J 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 This feature serves as an internal control for quality assurance and allows you to see deviations in your controls This flag is used for all applications We recommend that you run the control sample at least once for every panel Not editable OVL Overlap A flag Pass Ii or Check A is automatically displayed when a peak in the overlapped region is called twice It is possible to have two allele size ranges that overlap This feature serves as a warning for an allele calling error This is used for HID only NB Narrow Bin A flag Pass J or Check A is automatically displayed when a peak i
274. y Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Genotypes View 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 w GeneMapper Project Casework Project Eil it Analysis View Tools Help dst BE MuU We ws Table Setting Applied Biosystems Applied B z l Samples Genotypes Single sar ri a Marker Sample Name Panel Marker Dye Allele 1 Allele2 Size1 Size2 Height 1 Height 2 Peak Ar Peak An Mutatior Mutatior AEH Co AEHCo OS ADO AEH BIN 1 CO_Control Ofiler_v3 58 B 14 15 119 5 YM 2 CO_Control COfiler_v3 D168539 B 267 34 1487 0 1577 0 11898 0 12672 0 B a 3 CO_Control fsa CO_Control COfiler_v3 AMEL G x 103 41 2383 0 16110 0 Genem a 4 CO_Control fsa CO_Control COfiler_v3 THO1 Go ig OL 179 16 186 15 1077 0 1030 0 6695 0 6779 0 GeneMz GeneMs i ry 5 CO_Control fsa CO_Control COfiler_v3 TPOX G 8 223 0 1823 0 14283 0 Gene i i 6 CO_Control fsa CO_Control COfiler_v3 CSFIPO G 10 12 295 98 304 51 1249 0 1118 0 10362 0 9350 0 GeneMe GeneM i _ 7 CO_Control fsa CO
275. y Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Parts of the Analysis Methods tab continued Item Description 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 Sets the analysis algorithm 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 methods 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 5 4 Using Analysis Methods Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com 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 custom
276. y 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 B 34 Software Genotyping Algorithms Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Local Southern The Local Southern method determines the sizes of fragments by using Method the reciprocal relationship between fragment length and mobility as described by E M Southern 1979 Best Fit 2nd Order Curve Ad 1 990206E 02 Al 2 075795E 01 A2 1 545595E 05 R 2 1 000 Size Calling Curve Local Southern Method T T T T T 7 T T T 1200 1400 1600 1800 2000 2200 2400 2600 2800 Data Point Note This graph was generated using GeneScan 3 7 1 software These results are similar to results obtained when you use GeneMapper software v3 0 The Equation L c m m0 LO The equation attempts to describe the reciprocal relationship between the mobility m and the length LO o
277. ypes View on tabs page C 6 4 Navigation The use of this pane is described in Navigating pane in the Project Window on page 3 7 3 6 Using the Project Window Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Parts of the main window continued Item Name Description 5 Samples view These columns are described in Samples View columns Columns on page C 3 6 Progress The progress of analysis and import export of indicator projects is shown in this indicator Navigating in the This section provides information on how to navigate as well as how to Project Window access different views for samples files Navigation in Samples and Genotypes View See page 3 7 Info Tab See page 3 8 Raw Data Tab See page 3 9 EPT Tab See page 3 10 Navigation in Samples and Genotypes Views 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 Project e Run folder e Sample Genotypes View Panels root level e Panels e Markers As soon as you click the tree view controller indicated in the figure below the tree view
278. ze ranges can be edited Bins can be edited 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 SNaPshot samples X Axis displayed in base pairs Marker Indicator and Marker Range displayed Bins displayed To edit marker size ranges for microsatellites only a Select Binning mode b Click the marker indicator of the marker to be edited A red indicator displays c 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 a There is no binset for the panel b The binset was deleted 7 26 Using Plot Windows Samples and Genotypes Artisan Technology Group Quality Instrumentation Guaranteed 888 88 SOURCE www artisantg com Item Description Add Allele Allows the user to add a new allele call to an unlabeled Ctrl L peak as follows a Select Peak Selection mode b Select an unlabeled peak using the left mouse button c Select the Add Allele menu item or right click mouse button to open the Add Allele dialog box

Applied Biosystems Prism 377 GeneMapper Software User Manual

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