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GeneMarker® HID User Manual

1. 115 PEOCEOUIE s ositesdatet tenta Im MERE UIDES RD REL dan EM dE esM ent mou dE Cid DU 115 Save gna PAE sai ot ttes dtt t intet dot pute Do o mU et 115 CHAPTER S ADDITIONAL TOOLS e eoe e ete domu Modo el eno ugue dite ee v Deve ee a 116 AUTOMATED CONTROL CONCORDANCE ventu drea erica vea due vu cs ue d egi he Wed esum uon ae 117 NEGATIVE CONTROL CONCORDANCE sic ras tus diate diane ned naan ve vx Eu Eee Eo dd tes Ue URS eed vee e AE Iron pde 118 FILENAIVIE GROUP EDITOR lt cxccccc netu oi3 ta i aston Eis otio e tt on p e eov L itd cus a t Eee LOAD 118 OUTPUT TRACE DAT Ac ccc 119 eee eet rr Ar cr 119 CONVERT TX TITO BINARY uos iunt sexo ue iust san ux trae Pe OUR seo teuEs Edu vu eos een Mateos dus Rua CUu san uu Cem nu se Scr dieu oos Su MEG 121 EXPORT ELECTROPRHEROGRA sia terris ce LUCULENTER UE 121 REPLICATE COMPARISON TOOL n 122 CHAPTER 10 USER ieee pes ura alee saci eaa VER Maa S PN a EY Eo M ed cas vanessa vega Sen Ua Saa vade Voa e Vea d 128 d det ce tec cot nie pute es Lote oL 129 PROCEDURES M n 129 SERAVIANAGER S eite EL LIS
2. 88 PROBABILITY OF INCLUDSION Pl esses 88 PROFILE COMPARISON VIEW eee ee ee ee rene nennen nennen 32 36 PROJECT COMMENTS 5 isses aae ah RENE ERR VENERE VEN 32 PROJECT COMPARISON 5 555552 vay R Ea VAYA Feu Ea 33 119 120 PROJECT COMPARISON 5 5 120 136 PROJECT OPTION SETTINGS eee ee eene eene nennen eee nenne one 32 PROJECT PANEL EVE REN 51 PROMEGA POWERPLEX ebay ens yen us 56 PUEEUP CORRECTION ERO REND 14 PULL UP CORRECTION Un GER 17 25 QUALITY 25 QUESTION MARK SYMBOL rre PVP E PEN EE 94 R RAW DATA ANALYSIS socnsnensovasonnsoveteredexecoeaset 16 RAW DATA FOLDER 5 enn aan wA SE PE aaa ew Ea eR Van Y aaa eS 21 RAW DATA MAIN 5 5 11 RAW DATA TRACES 5255591 sear anau QR SIR ERE Dhu DON RR a 11 RE ANALYZE INDIVIDUAL 5 5 19 RE ANALYZE WITH AUTO RUN eee eee eee eene ee ee ee eene nenne 19 RE ANALYZE WITH RUN WIZARD cccccccecececscscscscscscsccecscsces 19 RECOMBINATION VALUES cececscscccccscsccccscscscscccscscsccecscsces 98 RECORD
3. VE SP CENE RN apap E MPES 38 Table of Contents SIZE TEMPIUATIE EDITOR c 39 PEOCE OIC td a TF mdr Mesas uoa d 41 lecons Qna FUNCHON o toe oa ege versat a a sail ean m doa Rosso ud obtestor usce ASR 42 WIGETO EXD CCE sire 553 yan 43 SIZE CALIBRATION CHARTS ararnir sonnin a A a seestubesesscuid san nue ueadcecuer A E E senstubesonncued seneuwubadass 44 PrOcCed aT PEPER EPC MAT PORE a ttr LIE REN IT ee Re SEP Ei SEL te auf 46 kons GIG FUACTONS eem ab m A RSA A PN ons 47 TO EXD CCE T Ec 48 CHAPTER 5 PANEL EDITOR 5 3 dE esse ane esa ace we ei cue coca ece swans ode su dul ue Ed 50 OVERVIEW c 51 Proje PANE M EN rcr 51 Panel Temblidtes ocu tura nota cm tete Lob oa pn iol el Cr ML ed ru fr 51 52 aid d 52 ole Ry AC 0 een 55 PROCEDURE 5 5 56 Pre Demed Panel MO 56 Custom
4. 52 58 5 28 DELETE SIZE bs SAY UU E UIT AR VER 40 DELETE SIZE STANDARD ccccccsccececscscscscscscsccccscscscscscecs 40 42 DELETE UNDELETE ALLELE 00cccessccssccssccsccceccccscccssccsccesces 26 DESELECT NODE sis 93 FREE 25 yn pu Kuh Mau 22 DISABLED veta vata vega raga etu wal guck vaa Vau alta Vau Val Vas 31 5 5 44 5 94 DISPLAY SETTINGS isos sees vessucaaciesauesenesesiesevess seas cucasesasibeseees 29 DISTANCE KB 55 DYE COLOR 11 E 27 55 EDIT BINS oe ku vs Vau V ek VERE VER E VY Vau ven oak ae aEd 54 EDIT HISTORY ia sists deta aaa ana Rude 131 EDIT HISTORY LIST sas SAEVUS DARE RASA CREARE uS 131 EDIT INDIVIDUAL cesses vios Eua E Fou ava suu SR Pes aud aD us 94 113 EDIT MARKER Joao eru bvraes vao aee 53 EDIT MARKER BINS tu Vu Ea aus 54 EDIT PANEL n Vu ku iniu au tn pu Gia
5. Deducing Missing Parent Genotype The partial genotype of a missing parent is deduced based on the allele calls of the available parent and the child ren In the example below the father and three children genotypes were available The Missing Mother genotype is deduced based on the genotypes of children and father Deduced allele calls are indicated with underlines Question marks indicate allele calls that cannot be made with the available data 112 August 2013 Chapter 8 Relationship Testing Father Missing Mother D S1179 D21511 D75620 CSF1PO D351358 THO1 D13531 D165539 D251338 D195433 VWA TPOX D15551 AMEL D55618 FGA child 1 child 2 child 3 D851179 D21511 D75820 CSFIPO D351356 THO1 D13531 0165539 D251336 D195433 WA TPOX D18551 AMEL 0555818 FGA Editing Personal Information Add Edit Delete Individual If an individual is the first to be added to the Pedigree Tree a family must be designated for the person If the person is already added to the Pedigree Tree right click and select Edit Node to change that person s characteristics To delete an individual from the Pedigree Tree right click the node and select Delete Node Family Name Enter a name for the family in the free form text box The Family Name field will not appear after the first individual is added to the Pedigree Tree All subsequent individuals added will be considered members of the family
6. 42 NEW TEMPLATE isiesvneus sun uk eub su busub unus Eu beue pe rs 15 NUCLEOTIDE REPEAT sessissscteccsnsstesavevetesivevevessectevevsvase 52 54 NUCLEOTIDE REPEATS issues usen peek pas NE EVE PEDE 54 NUMBER OF LIABILITY CLASSES scccccccsccecscscscscscscscsccecscsces 98 O OFF LADDER PEAKS 4 OFF LADDER PEAKS osea eus au anao vu Va cn a auo ga va euo puse a TR ORE 63 OFFLINE REGISTRATION cccccccecscscscccscsccccscscscscscscsccecscscss 7 9 OFFSPRING 93 OL OFF LADDER PEDIR UE 26 ONE COLOR ANALYSIS ux ks paa ERRAT Es Uus 16 ONLINE REGISTRATION cccccccecscscscccccscsccscscscscscscsccecscscss 6 8 OPEN 28 34 OPEN DATA FILES siis ea ss ax ex Xe Eus EXE X Pa ex PR DEN 11 OPEN PEDIGREE FILE eee eee een 96 98 103 107 OPEN PROJECT sab Qux gua Fav vaa Dove ako Qu pu Viet alba Vus 28 74 er sje Lp 32 August 2013 Index OUTPUT TRACE DATA iva va uve ua iva VAN EAE NEN Y NN ua ME new d us 34 119 OVERLAY TRACE UT aS 52 OVERTIME PROTECTION SAFE ERES NOR RN TAX Ray 130 P PANES i Spiri Ge ueni 15 PANEL CREATION usse 56 PANEL EDITOR 15 16 18 19 26 33 50 51 52 56 57 58 59 61 67 70 74 105 7 5512 NN Lt de
7. 33 5 98 103 107 PEDIGREE TOOL qut cave d VE Desa si 93 PEDIGREE TREE E Sdn E NE SEN MENTI VE UN VA TUS LUE ad 93 PENETRANCES aeu 2 auk a ERA 98 PERCENTAGE vos visis ex av X ia GE VY EX UE ME RAE os 18 PL BEYOND PIOIDY irai bas ERXEE TA ER RERCSU SAPE UP RNP EIE 26 POSITIVE CONTROL 552 25 sax Eua 30 POSITIVE CONTROL 33 117 PRE FORMAT CET E Talis 96 PRE DEFINED PANELS cis stssncnsssencsnuvenses pusRREPERESEEEEEREONRE ERE 56 PRE DEFINED SIZE 5 41 PREFERENCES BOX nua a Va UR Maia wu uia du Red 29 PREPROCESS RAW DATA 652a tcanucaeiearncaetcees 47 PRINT 5515533 PRESE LEER DNE AN Fe EXER AN SR EY UR 73 PRINT ALLELES aaa ua uU e VE E 72 PRINT MARKERS 5 isa e des eane essa EkxaPExesiX aba vex Pul ed exe sa e Pagos 72 PRINT PAGE SETUP dese e ve dn ao ann ko sotoko 73 5 72 PRINT REPORT 22 29 32 34 36 66 71 72 73 76 5 72 5
8. 130 RECOVER OED VALUE oscievsdswsdesdesvddestsvsdeovsdoabseverevecesesexas 131 REFRESH PEDIGREE e eee ee ee ee eee ee eee eee ene enun 98 103 107 REGISTRATION U UE 6 REGISTRATION ID Far a ad 7 9 RELATIONSHIP TESTING ssi renun ERR yao SF eX en Ren 93 RELATIONSHIP TESTING scsscsvevcsovessusvessvnseesesvestvoesssesusesvessbe 32 RELOAD PANEL 2 Cueva Qut Y va Qu edo va pata va du da ua 52 RELOAD SIZE STANDARD Leve ku n hi nin nh nin Un nnn n ao uin oo ku pud vu naga p 40 REOPEN PROJECT vs va epi 74 RE OPEN PROJECT 28 REPLICATE COMPARISON sccecscscscccccscsccccscscscccscscscsccecscscss 33 122 REPORT SEELTHNGS 35 REPORT SETTINGS ICON isi vsus aee yRvE RE ARE RNAs a ERERER EY AY Se ER VR EE OY 27 REPORT SETTINGS PREFERENCES TAB eee eee ee eene nne 31 REPORT STY E esxeseke pete Vu a ce gae ve teu te va ava eg 67 REPORT STYLE OPTIONS i ies nena nra rn n gun pu Eug aun 27 REPORT TABLE baud bad 27 67 REPORT TABLE LAYOUT iiirved soe usur ene Sem edu 67 REPORT TABLE ORIENTATION eee ee ee eene eene eee nete nenne ene 67 RESIDUAL
9. 54 SYNTHETIC GEL IMAGE eee eee ee ee ee ee eene eene eene eene enun 23 SYSTEM REQUIREMENTS sve p oa pen RFRER ERE 5 T TECHNICAL SUPPORT iussis Ey a EE 34 TEMPLATE NAME i ve is av a da an po no au eanet 15 TRACE MODE 5565avsta gs t arut ass aaro ap 60 TRACE 60 5 UPDATE CALIBRATION ies eoa a Futsal ausa vovv e vas vaa apa ad 46 USB KEY o ue Deva Dentes Due ta va aeu Deve Da Va eua 5 6 8 USB PORT conu uva ntu Eu aya o ER 5 6 7 8 9 USE LAST TEMPLATE cccccccecscscscscccscsccccscscscscscscscsecscscscscecs 15 USE SIZE STRING FOR LABEL eee ee eee eee ee ee ee eene nope een 29 USERID oe RASA 2IEAE SERA UFU U FED CE edes 6 9 USER MANAGEMENT ee eee eee eee eene nennen eren 34 129 USER MANAGER HISTORY esee 130 USER MANAGER SETTINGS issii Se ERAN PRERERX SERES CRX FX ej 130 V VERTICAL REPORT TABLE ccccccccsccccscscscccccscsccccscscccscscscsecs 67 VIEW HISTORY seta sen auta aka o2 UTR MUN aluo 27 131 VIEW MODE cs ea e 47 VIRTUAL 55 VIRTUAL PANELS 59 VPANEL PANELNAME e eee ee re oru ne re ene ee oe opo eu penu neuere Een 59 August 2013 Index
10. gi i di Tal Monee aros m Ore Cobo Ho Make Wan JH TM 4 ence HIM Minor EM Tea Are 09510179 nu Tha amo aon qo y ro 41 oer r Mb eae m n gt 03511793 a 14 14 nea now ne Mann ota Hi 2 m armas 2 HR i Tivab i wane Lo 3537 t nta 03579 faa ti TRE CTS Wet cd wee Li i DEO ee tone te Dhin aa Aer Da 3i m m Lg WE X e Samy uaan 0 ES an aru 16h Ph 876 alt nn an 2 a ernro nme 213 730 anes 4 1 csrim 710 uu o 0X ow an Em 5 Cm ma a aS 3 1 joi 5105 518 Om THON 753310 oar a 789 simis Ee QUEM Se DSa 43124 ee 3 ia ET 8 11 Tum 33 910 os Oos Q psss Sed hes mi um 2 ILS LEE pi er 3 He m OTP ZL 01937 1524 1721 jen lud d Osan 11514 Dasi 1214 nm 19 1 NS n 1 24 f a 2 AA 516 19 diaa 00721 1613 5 944 5 APTE 1 TP asian OC 3 Nw 3 I ad ESI EM milia man 42159 ps TAL 1287 513 10 3 Dr45433 7473s Eu Wr ae ey 3 E Hs ee a psssme 510211 imam 673 14A T um 202124 Mm pum 22 NM men tae Dress TAL 1215 100 z 1I 7 ave 4 2 AMEL KY 0s tb O63 Lus 18 1 Deme noi gs Uns Ux T 1
11. Gr PUE EK EO ED FUR 72 CHART OVERLAY eckxsx vue bx Fa Ex Ix E RES FEAR EE E EXER FOE PESE 72 CHART SETTINGS even icr dc M EIE MUN 29 CHART SYNCHRONIZE educa o OE UE 47 CHECK RANGE IN eura cass sev Eee rex us 60 177 eR n 95 Qf ccr C TT ppp 29 CLIENT COMPUTER rus Era 7 CLOSE ALL RC HORE 29 Gu QNT M 32 74 o pede 32 74 32 74 COLOR MATRIK RR 14 COMBINED PROBABILITY CPI 2sssccessoeccessceccescceccesccecceccs 88 COMBINED PROBABILITY OF EXCLUSION CPE 88 18 essi 28 26 el rimi 9 CONTROL 116 117 CONTROL GENE iiv OE Er EVER E renee 55 CONTROL IDENTIFICATION ee ee ee ee ee eene none nenne 118 119 CONTROL DENTIFIER xsssseestses sesisesxessssus ve veuve se vexeusteecz eres 119 CONTROL 119 CONVERT IMACINTOSH 5 5 CONVERT TEXT TO 121 CONVERT TEXT TO BINARY FILES eee ee ee ee ee
12. 87 RIVINE 88 RUN mre 31 34 RUN METHOD 4s a REUS EE 29 RUN PROJECT i55 15 34 RUN USER PROTECTION cccececscscscccscscsccccscscscscccscsececscsces 129 RUN 15 29 RUN WIZARD ADDITIONAL 5 5 18 55 16 RUN WIZARD LOAD DEFAULT scccccccccsececscscscccscscscsccecscsces 18 RUN WIZARD SAVE DEFAULT eee eee eene eee eere eene neon 18 RUN WIZARD TEMPLATE 5 15 August 2013 Index S SAMPLE FILE TREE esee es enean ea aaa o eu va va vn eu a va Va e a a gea ok uas 21 SAMPLE GROUPING s v o vus yu sopa nu s uhu oe sus ia anu naui 23 SAMPEE TES oar aeons 41 45 SAMPLE INFORMATION ccecscscccccsccccccscscscscccscsccccscscscscscecs 22 SAMPLE SCORE scesero eaae 44 5 5 22 5 22 SATURATED 14 SATURATION CORRECTION 1 555 saeua agua NEG Fa Fa EE 12 SAVE AS NEW PANEL eut a en aa ena vaga ena egeta rub eU Vs 58 SAVE AS NEW SIZE STANDARD cc
13. Allele Comments Right click an allele in the Electropherogram or Peak Table and select Edit Comments The Edit Allele Comments box appears Select a comment from the j c Comments Template list or enter a new comment in the Comments field Click OK and the comment will appear in the Comments column of the Peak Table Only one user edited comment can be added to a peak Comments automatically generated by the software cannot be removed Additional user comments will simply be added next to the software comment owen View History Opens the Show Edit History window Shows a record of all manual edits performed on the peak The Show Edit History window is only active when the Help User Management Settings Record Data Edit History option is selected Print preview and print or save as jpeg png or pdf for paperless record of audit trail See Chapter 9 User Management Report Table The Main Analysis window Report Table contains additional information about sample peaks See Chapter 6 Reports and Printing Navigation The Report Table is linked to the other frames in the Main Analysis window Double click on the desired allele OR use the Arrow keys to move to the cell of interest and hit Enter key OR use Alt Arrow keys to move to different cells and zoom in on the peak in the Electropherogram Select multiple cells by holding down SHIFT key OR hold left mouse button and drag over desired cells The rules by which t
14. 5 53 MINIMUM MBALANCE eee eee eee eere neenon ener rete tese sone 53 60 hu EP I CM 72 32 76 79 MIXTURE ANALYSIS 81 5 87 MIXTURE ANALYSIS LIKELIHOOD 5 89 MIXTURE ANALYSIS OVERVIEW cccccsccccscsccscscsccscsccscscsccscsoss 77 MIXTURE ANALYSIS PROCEDURE eese eese esee ee eese eese 6 6 55 77 MIXTURE ANALYSIS RESULTS cccsccscsccscscsccscscsccscsccscscsccscsoss 80 IVIODIFY SIZE STANDARD eee ee ee ee ee eene ne nene hehehe teneo nene 41 MOTHER ia sehn deu dut a auta Ue M 94 114 N NAVIGATION seta rota Fete en aun 23 NEGATIVE CONTROL 5s izeissvussvuucosdvexseosuFoFarekUs ose cFVKERVRERE 30 NEGATIVE CONTROL 118 NEW FAMILY 97 NEW INDIVIDUAL saxa andata auk n unn nau 97 NEW PAGE FOR 72 NEW PEDIGREE FILE 5 suse s ea yov asa sare FEE ka eas 98 103 107 NEW SIZE STANDARD
15. g gos g o o E s go go Jo gs E os F o Hide Extra Sample Names When data is displayed in Vertical Orientation the sample names are repeated for each row of data that the sample is associated with If Hide Extra Sample Names is selected then the sample name will only appear once in the first of the rows it is associated with Forensics The Forensics Report Style displays the Quality rank and Allele Label of each called peak Samples are listed in rows in the far left column and Panel Marker names indicate the columns at the top of the table Allele Report Settings Options Report Style C Allele List V Extend Diploid Homozygous Forensics is the default Report Style in GeneMarker HID i Forensics NOTE Forensics requires that a Panel is applied to the data See Chapter rds 5 Panel Editor Peak Table F M Allele Count pm Features Options Sample Name FileName 2 h llele call Extend Diploid Homozygous Repeats the same Allele Label in the second Orientation P undas Show Only Uncertain Alleles Horizontal Vertical allele position of the marker when only one peak is detected in the marker Only active when the Edit Panel Ploidy option is set to 2 Diploid Show Allele Name Size 0 1bp Height Area Allele Name is displayed in the Report Table by default regardless of table Orientati
16. n saa zen ana naan ae ees 5 X IE UL Si Fie Name 1120206 20 001 561 12 12070820 512 042561 Diess99 12 mon mnnnnnnnniu Other Features and Considerations Disable Entire Replicates The replicate comparison tool allows the user to remove entire replicate groups from the comparison To do this simply right click on the Sample Name column of a replicate group and select Disable Allele and status calls of disabled replicates will be colored dark grey Simply right click on the Sample Name column and click Enable to undo these effects Disabled replicates are never exported regardless of which reporting option is selected more Bi Ho Sampie Hama LE 1 ENDE eT og NM 125 August 2013 Chapter 10 User Management Disabling Allelic Ladders Because the report table expands to include every allele call for a given replicate group it is recommended that Allelic ladders be disabled prior to entering the tool Otherwise the report table will expand to include every ladder peak making analysis of relevant peaks more difficult To disable ladder samples in the Main Analysis Window simply right click on a ladder sample and select Disable NOTE You may be prompted to re process your samples If the disabled ladder was used in your analysis procedure select No File grouping must be done after allelic ladder samples have been disabled Otherwise the ladders will
17. 21A um 75 a e Comme MDC etm Cini ntc Korm sin ill pepe Compared ls Ama Find Fun Sm gl he eee er hon dare aan persons ns stades dos oar hel I Mmi mime Ma Mapa m dli jiro Dmi EA Contesting contributor two as a contributor to the mixture m Meer ee Tie pua Finn m 115334 M m nt san Dots 3 3i1 02 ASA 65431 3031 AR nom DEM samy Ust UR 30 aru n 4 cosmo 101211 04553 5 3213 730 14 5 Oma 151 0 51 OFM 1516 15 5004 THOI 783310 al 183 715 41 86M 423100 QM m 1214 58 15 E Sri Wit 1361 Fa Drs ea 01430 14 ma Yn 121114 D 1714 n 4515 15 16 19 113 092721 1813 5 22100 asin nsum nowe5 8 ETT 545 t3 Dt850 121532 18 nixa MB 1215 mas n n xv x lt r m osse apid iam mar 471 m 202124 1579 pann 23 4 2024 530 Cima om 1 1 DET IE x2 aq 1 a Cammen Cordt Harri ctm rod 9 Cinisi TM hypotheses vo fuoatrege thal ve qoi e Rom coniute 2 33 4neenm Door nt fem ed to congu 1 Uritu TT Tam ereer Mapex Meg canana Ame Mec ro Omir iA SEE RMWE TELLY 85 August 2013 Chapter 7 Mixture Analysis Contesting contributor 1 as a contributor to the mixture
18. Edit Individual Person Info Name Free form text box to enter a name for the individual Display the individual s name in the Pedigree Tree by clicking the Show Individual ID icon in the toolbar Gender Select either Male Female or Unknown gender for the individual Male nodes are squares female nodes are circles and Unknown gender nodes are displayed as a circle within a square 113 August 2013 Individual Info Name 3 Render C Male 4 Female Unknown Father Mother Deceased Population Benotypes M Affect Status Unknown C Unaffected Affected Conteste n From Sample File PAT_1_M tsa C From Database None Manually Input e Cancel Chapter 8 Relationship Testing Father Mother When more than one mate is displayed for a Mother or Father a drop down menu allows the user to choose which possible parent to associate with the child NOTE The Gender and Father Mother options are not available when adding a mate Affected Status Affected Status options are available to mark individual nodes for genetic linkage calculations Before marking individual nodes with Affected Status click the Pedigree Parameters icon in the toolbar and adjust settings accordingly Unknown The individual s phenotype is unknown The node is displayed as an empty square or circle Unaffected The individual does not show signs o
19. General Settings Show Navigator When selected the Sample File Tree will automatically be displayed in the Main Analysis window after data processing Show Gel Image When selected the Synthetic Gel Image will automatically be displayed in the Main An window after data processing Show Report When selected the Report Table will automatically be displayed in the Main Analysis window after data processing Display Settings The Display Settings tab is used to set how the data is displayed in the electropherograms Allele Label Decimal Precision Select 0 to 2 decimal places for peak size labeling Mark Off Allele as OL Select this option to label alleles that are outside of allele ranges as OL Use Size String for Label Select this option to label peaks in the electropherograms according to size instead of the allele label To display a rounded size string set the Decimal LL e Precision to 0 SEA Bingley Sets Foeeric Sefingi Uter Larger Font Doubles the font size of the allele label characters This fpe Fed Lobel increased font size will carry over to the Print Report Demim x Hed pe UH ubl UL ara Soren Flag Variant Alleles in Ladder Select this option to flag peaks detected 4 rint in variant allele bins of allelic ladders E RH Jaiei eM Show AII Allele Labels Will display all allele labels when all dyes are disp
20. Corte ABUUT US ied BB Conil ARNM h hid D EUG Corta ABUUT 00 FO Conhol 07 hid Consol 007 b 06 h Us_Contial UO c 05 ladder All l hk dtd m Control ABO OSI 15 5 FG xc at Ft ik HU a ci Ee 5 702 Seateol 007 zm PES Pr Pr wp vt z f 007 zd pisces y 5 ipo Sree UST 0165539 C xy1 0 IPOX g rj a i a g g i g B a s a a a a a a 63 August 2013 Chapter 5 Panel Editor File View Project Applications Tools Help xm S 9 JAE A Make s 5 Untitled gt H A Raw Data 3 allelic ladder 1b D8 hid Allelic Ladder H 6 Allele Call 2 2 B EO4 Control ABOO1 b O5 hid 2 y Control c O5 hid 3 Control 007 06 hid B B LO Hf allelic ladder 1a 08 hid 5 LD H02 allelic ladder 26 D6 hid 6 y LD H03 allelic ladder 1b D381358 50 100 120 15 121 1 17 129 1 F 2 Control 007_a_O6 hid Ref Ladder 3 allelic ladder 1b 08 hid 0351358 80 100 120 140 4 Control 007_b_O6 hid Ref Ladder HO3 allelic ladder 1b O8 hid 80 100 120 140 160 180 200 Page Down Alternative work flow Select the desired panel from the drop down list in the analysis settings to use one panel for all samples in a project When this workflow
21. De amp Cancel This feature is used only in cases where the data is extremely difficult to analyze and cannot be corrected with the Smooth function Peak Saturation The software will analyze saturated data points by creating a synthetic estimate of the peak shape based on the curves prior to saturation The results will be less accurate than that of non saturated peaks Baseline Subtraction This function removes the baseline completely so that the Y axis will be raised above the noise level It uses 20 of lowest intensities to the right of the beginning of the range and looks at the trace in 500 600 frame sections 16 August 2013 Chapter 2 General Procedure Enhanced Baseline Subtraction This feature is used only in cases where the data has excessive baseline in one or more of the dyes or has an interfering slope from the ion front in the smaller marker ranges The function uses the second derivative of the absolute value for every 30 data points and looks at the trace in 300 frame sections In situations where there is an extended ion front in the mini STR range Enhanced Baseline Subtraction should be used Pull up Correction This function removes peaks caused by wavelength bleed through to other wavelengths The function should be disabled if a Manual Pull up Correction was used in the Raw Data Analysis window Spike Removal Removes peaks from voltage spikes caused by micro air bubbles or debris
22. GeneMarker HID STR Human Identity Software User Manual SOFI GENE rics Software PowerTools for Genetic Analysis www Softgenetics com Copyright Licenses and Trademarks 2001 2012 SoftGenetics LLC All rights reserv ed No part of this publication may be reproduced transmitted transcribed or translated into any language in any form by any means without the written perm ission of SoftGenetics LLC The software is copyrighted and cannot be altered or given to a third party without the written authorization from Soft Genetics LLC Thes oftware may belicen sed from SoftGeneticsLLC Mutation Explorer Mutation Surveyor JelMarker ChimerMarker NextGENe and GeneMarker are trademarks of SoftGenetics LLC All other product names and or logos are trademarks of their respective owners Limited Liability of Using the Software In no event shall SoftGenetics LLC be liable for direct indirect incidental special exemplary or consequential dam ages including but not lim ited to procurem ent of substitute goods or services loss of use data or profit s or business interruption however caused and any theory of liability wh ether in contract strict liability or tort including negligence or otherwise arising in any way out of use of this software even if advised of the possibility of such damage SoftGenetics End User License Agreement GeneMarker GeneMarkerHID JelMarker and Mutation Surveyor NOTICE TO USER PLEASE REA
23. GlobalFiler allele frequency table is populated from the marker information of the Identifiler and ESX publications Fusion Butler J M Hill C R Coble M D 2012 Variability of new STR loci and kits in U S population groups Profiles in DNA Identifiler 15 2003 Butler et al John M Butler Ph D Richard Schoske M A Peter M Vallone Ph D Janette W Redman and Margaret C Kline M S Allele Frequencies for 15 Autosomal STR Loci on U S Caucasian African American and Hispanic MG July 2003 Vol 48 4 Paper ID JFS2003045 484 htt www cstl nist gov biotech strbase pub pres Butler2003a pdf New Pedigree File Select New Pedigree File to create a new pedigree with multiple families OR select New Family to add a family to the pedigree file Enter the first family member s information into the New Family New Individual box and click OK to create a new Pedigree Tree E Open Pedigree File Launches the Load Pedigree File box Select a PED or PRE file to upload the SMP file will automatically upload and click OK Save Pedigree File Launches the Save Pedigree File box Enter filename and change directory to save the Pedigree Files PRE SMP DAT Show Individual Name When selected the individual ID will be displayed in the nodes of the Pedigree Tree Update Sample Data Select to refresh the Mendelian inheritance calculation after a node or allele is edited and after selection a different family when the
24. LLLI PELA 129 FST OR E A 130 ai saree ucc secula 130 EDIT HISTORY ADDIT ERAI toes tede detta e tee aeu es du v cda nite c I Me dE 131 INDEX ME S 133 3 August 2013 Chapter 1 Installing GeneMarker HID Chapter 1 Installing GeneMarker HID Chapter1 Installing GeneMarker HID Computer System Requirements Local Licensing Option Network Licensing Option Questions 4 August 2013 Chapter 1 Installing GeneMarker HID Computer System Requirements GeneMarker HID software has been tested and validated for various computer systems The minimum system requirements are Windows PC OS Windows XP Vista Windows 7 Processor Pentium III 1 GHz CPU RAM 512MB Available hard disk space 20GB Intel Powered Macintosh Parallels desktop for Mac Mac OS virtual machine dependent or Apple Boot Camp or VMware Fusion Mac OS virtual machine dependent RAM 2GB Available hard disk space 20GB Installation of GeneMarker HID is not supported on Linux or UNIX based operating systems GeneMarker HID will only recognize PC file formats To convert Macintosh file formats to PC file formats please download the ABI PRISM 3100 Genetic Analyzer Conversion Utilities to co
25. jpo Page Setup Opens the Page Setup box Choose the paper size margins and orientation Portrait or Landscape Content Options Opens the Print Report options box See the section above Report Content Options T Jo Zoom to Fit Zooms out to view the entire Print Preview page Zoom to Width Zooms in to view the Print Preview page at maximum width without losing information off the screen T Zoom Ratio Enter percentage numbers to increase or decrease the zoom aspect of the Print Preview page c 73 August 2013 Chapter 6 Reports and Printing CODIS Report Tools Export CODIS The CODIS Report feature allows users to create an mir osan r CHF OC hal exportable report file for easy input into the CODIS database E The CODIS Export window will appear with several oa f z 8 iipon Paich ideraser options for modifying the header and classifying the samples To change the classification information for the samples simply click the cell you wish to modify and select from the options in the drop down menu Comets D8S1173 IDAST Once you ve updated the information click OK and the option to save the CODIS file will appear Select a location and save as either a CMF1 0 dat file CMF3 0 xml or CMF3 2 xml As per US DOJ CODIS Interface Specification CMF 3 0 CONTRACT NO ITOP 97 0087 Sub Task Order 26CODIS and Interface Specification CMF 3 2 Revision 9 DOJ FB
26. 054 4231 0 55 D351358 15 Check na 972 100 7736 1 00 D351358 16 Pass Example for D3s1358 87 August 2013 Chapter 7 Mixture Analysis Observed proportions 14 0 1237 1 Mx 2 0 14 15 0 1897 1 Mx 2 0 14 16 0 3505 Mx 2 0 36 17 0 3361 Mx 2 0 36 Residual 0 0034 Heterozygous Imbalance HIM Example for D3s1358 Major HIM 932 972 0 96 Minor HIM 343 526 0 65 Expected proportions The four allele model where Mx 15 Combination AB CD AC BD AD BC BC AD BD AC CD AB Allele A Mx 2 My 2 Mix 2 1 Mx 2 1 Afx 2 The three allele mode Combination AA BC BB AC CC AB AB AC BC AC BC AA AC BB AB CC AC AB AC BC BC AB Dwo allele model Combin AA AB AB AB XA BB AB AA BB AA AB BB BB AB ation the proportion of sample from the first indrvidual B Mx 2 1 2 1 Mx 2 Mx 2 1 Afy 2 Exclusion Probability for mixture profiles uses the allele frequency of a given population for each allele in a mixture sample GeneMarker HID calculates exclusion probabilities for two person mixtures and mixtures of three or more contributors When the CPE is multiplied by 100 RMNE Random man not excluded it provides information about the percent of people in the population that are excluded from being contributors of the mixture because they have one or more alleles that are not present in the mixture Probability of Inclusion PI
27. 3 Any negative control sample that has peak s will result in a negative control NC error in the Project Summary Bar 500 PC error 0 1 NC error 1 1 Ladder error 0 1 Failed 0 Filename Group Editor Project Apply Sample Grouping OR Tools File Name Group Tool The Filename Group Editor can be used to group family members or other related samples in the dataset based on their filenames for simplified analysis SF File Name Group Editor E Procedure 1 Select Project gt Apply Sample Grouping cm 2 The File Name Group Editor window appears b s 3 Click the Load Files icon and select all files to pair if Bt the dataset samples do not automatically appear in the mre REET ha File Name List field are 4 Choose Match by Sections or Match by Fixed Position T 5 Enter values for the Group Identification and Control m Identification fields 6 Enter a Control Identifier value and click Match 7 The samples from the File Name List will be paired into PrE z Vigo cm groups in the Matched Groups window Rie 3 Ceu as 8 When the samples are grouped correctly click OK OR Maleh to idee by Sestian F IY ucc click the Save Groups to File icon to save the grouping information as a tab delimited Text file 9 The File Name Group Editor window will close and the grouping information will appear next to the sample filenames in the Sample File Tree
28. Panel Editor Action Help box appears Pate eee ae te a Delete Alleles Hold Oal key hold left mouse and move the mouse to select a region dan as a cyan shadow en nihi bk mt 6e mome to delete aletes covered The markers Gat me covered wil be erased as well Toolbar Icons Panel Editor Found in the Run Wizard Template Selection box OR the Tools menu Create New Panel Opens the Create New Panel box Follow the steps in the Create a Custom Panel section above Save Changes Permanently saves Panel edits to the currently opened Panel file which is located in the SoftGenetics GeneMarker HID Panel directory for Save Changes with Signal Info Permanently saves all Panel edits including height information which is used with the Major Panel Adjustment feature 59 August 2013 Chapter 5 Panel Editor NOTE A Panel must be correctly aligned with peaks in the dataset before selecting Save Changes with Signal Info in order for the AutoPanel Adjust in the RunWizard and the Major Panel Adjustment features to work correctly gt Delete Current Panel Marker Deletes whichever Panel or Marker is currently highlighted in the Panel List This action is irreversible Show Dye Allows the user to select a single dye color to view in the Overlay View Cycle through the colors by left clicking on the icon H Trace Mode Single left click to cycle through the options or use the drop down menu Trace Overlay di
29. The summary bar at the bottom of the main analysis screen displays information about the results Ladder errors may be flagged with a yellow vertical bar and green allele label to alert the analyst if a non control minor peak is higher than expected figure 1 If any peaks are not in the expected bin they are flagged with red allele labels and red vertical bars figure 2 The analyst can return to the panel editor to determine if there were problems with the capillary for that particular ladder figure 3 GeneMarker HID Untitled File View Project Applications Tools Help gt R61 ETSI pE em E A a x 1 1 Ep LA Marker None 551 Untitled Co Raw Data Gel Image amp Allele Call Ea PC positive control fea LD identifier LADDER fs Moe MIPZOScasel eviden M gt MIS aset spect fs MIOS bassi Vlt aa 100 150 Identifiler LADDER tea AMEL D55818 150 100 am ae Page tip Page Down 26 2 20 2 48 2 New 6 samples C error NC error O 1 Ladder error 1 1 FFatled O Flaggeds 3 200 2 400 X T s F 2 200 2 000 1 800 1 600 1 400 1 200 1 000 800 800 400 200 o Figure 1 Yellow and green flagging alerts the analyst that although the panel is aligned with the ladder bins some of the minor peaks in this ladder have a higher RFU than expected 61 August 2013 Chapter 5 Panel Editor adders _bad SGF E E
30. and apparent stutter product above the normally expected range From the Main Analysis Screen select File 2Open data or Open project 2Add POkay Launch Run Wizard from pop up menu or from Project drop down menu Select Appropriate Template and Run Parameters see chapter 3 Select Mixture Analysis in third screen of the Run Wizard Enter desired values for Valid Mixture Peak Percentage and Min Number of Peaks Marker 3 is appropriate for a mixture from two contributors pte e qe x 6 Review allele calls and summary information 77 August 2013 Chapter 7 Mixture Analysis Review Results Mi GeneMarker HID altered file mimic suspect SGF File View Project Applications Tools Help Gb Ee m D amp 8 bx KS 1 4 l Marker None Sa gt altered file mimic sus a Gu 9 Help is z bi Gel Image arker Allele Aillele Allele A11e1e 7 LD Identifier 15 X 16 OOo o NC piss presses 2 B MX MiXO5ca eae 2 Bg MixOScasel oo pasme X 2 s sss s x ec a as x x jesse s x n X c E 3 me Xn Eod wa esses s E a s X x eese s x s X occ E 6 120 140 160 180 200 220 240 260 280 300 320 340 MIx lt O05casel_evidence fsa Mixture x 120 140 160 180 200 220 240 260 280 300 320 340 Size Height Ht_Ratio Area Marker Allele Difference Quality Score Comments Quality Rea 1248 3481 0 53 20866 D351358
31. 1 2 71 Li hl arker Marne 2008 11 03_CO4_Ladder_15 50 06 fsa peste esa orssz 150 2008 11 03_ Ant LADDER 14 12z 17 fsa 356 SPS x osm AAE csFiro 150 250 H H Figure 2 One of the ladder samples in this run has peaks that are out of the bins These out of bin peaks are flagged with red allele label and red vertical bars Often on full CE runs there is slight migration variation from beginning to the end of the run In the following images we see several allelic ladders that migrated more quickly than the others The auto panel adjust and auto select best ladder parameters provide the needed minor shift to align bins including variant or virtual bins A list of all passing ladders is located in the Project Panle list under the panel Name Select the allelic ladder from this list to highlight it in the electropherogram and see the bin alignment 3 B Proe Panel Glsb llle Peet Globe Panes GF Globale Pas GF BT Globe Paret HOT isdi Globales Panet HUZ todd Globale Parcel akeke inl Globsitie Ponet HOA abio lead Globale Fare HUS ale inde 3 1 Globsiier Pace iii T A 23 fae a Let Flange A gen nm Cori Detonce feb Comment 050 25x 0 8 9 0 nn 4 i D 1 En 250 1
32. 10 The information in the Pedigree and SMP files will recreate the Pedigree Tree and associate the nodes with the samples in the dataset Create an SMP File from a PED PRE File If an SMP file does not exist for a Pedigree File or has been lost a new SMP file can be generated with the Pedigree File Name Match Input PED PRE File IC Users SoftGenetics Desktop T estPed pre Open EMP File 3 tool Input Filenames 11 jenetics Desktop TestPed 279878 20_04 fsa Add 1 In the Main Analysis window menu bar select Tools Pedigree File Name Match Family Identifier from character 1 ta in file name The SMP File box appears Individual Identifier from character 7 to 3 in filename Click the Open button s od PUR Select a previously created Pedigree File PED or PRE I Extract sting within senerators Click the Add button Select all the samples included in the Pedigree File ziv JU xus Click the Process button The SMP Table box will appear with filenames aligned in family groups Click the Save As button 0 Save the SMP file to the same directory that contains the Pedigree File 11 Proceed through steps 1 10 above POON ON O2 N 96 August 2013 Chapter 8 Relationship Testing Create New Pedigree 1 In Pedigree tool click the New Pedigree File icon 2 The New Family New Individual box appears 3 Enter a Family Name the Individuals Name Gender and the Sample File to as
33. Dara Process HID Analysis uid Ga Dokta Run Wizard Data Process Dd a enisi Procedure hainn hen il 5 The Data Process window of Run Wizard appears skim 8 Ered ymo sg IJ moni rrerced Soon 6 Select the appropriate analysis settings in the Data Process F Peak Suuaion Batalne window and click Next to continue Sici fF PulupComechon E Sams Cu eee gm Icons and Functions Raw Data Analysis Loris ave Derma Auto Range frame The range in camera frames will automatically find the processable data range If Auto Range is not selected manually enter the start and end frame numbers of the data set for analysis NOTE If automatic size call fails due to high saturation de select Auto Range and manually input the required data range tal Intensity Coefficients Allows for manual correction of excessive bleed through peaks best used for experiments with one color analysis Allows for manual correction of low RFU by using an number greater than 1 to increase the RFU Smooth Smoothes the baseline by eliminating smaller noise peaks Enhanced Smooth I Tl Alis Ll Aars bns jo si Faai Duetechen intend TET i irita xXx 5 1 ja Global Mug i Pak fime y Data Intensity Correction Dolntensity Correction Coefficients Det Dez Dyes Det 1
34. Entera Panel name in the Name field 3 Choose the appropriate Analysis Type from the Type drop down menu 4 Select Manually Create 5 When finished click OK 6 7 Create a Marker in Panel Editor Panel Editor eS File Tools Help B BH x Era Se 3 Hep d 1 PowerPlex 16 Test Panel Eg PowerPlex_16_4DJ af PowerPlex_ES af PowerPlex_Y 7 9f Profiler 1 400 Y 9f Profiler_Plus 1 200 F Profiler Plus CODIS Coe EE iti iti iti DERE iti DERE ERE ERE REF GE B ore je S 4 ae 600 i 2 T Create Marker 7 SENE PUE All 400 Change Marker 1 Test Panel 200 LA Delete Alleles is din ente Plus LADDER fsa Bj SGMPlus MixD 5casel evi 131 5 763 B SGMPlus Mb4DScsse vic Due Marker Sie Left Range Right Range Aele Name Conil Distance kb B SGMPlus_MlsO05case2_evi B SGMPlus_negative control B SGMPlus positive control fs Panel Name Iest Panel Ploidy 10 Adjusting Panels Manual and Automated Panel Calibration It is common for panel alignment to be shifted due to variations in genetic analyzers or run conditions such as temperature injection time Markers or bins can be manually aligned to the allele ladder using the shift and mouse key or the panel can be automatically calibrated using the auto select best ladder and auto panel adjust in 57
35. Ev alain NEHE PETA ESTN Pak Toue Wap pa mere Ru poate m Mette POS Poste Contro Temexate Eniter m Co ae ee e i eee El nu me ee Paniren Coriri SSmi mmm map v te ad Datel i Gengas 3 M ilete 1 Arie impe Genotyeea h m Samot 1 12 13 i b2t511 1 Draam n E corro 1 5 353 Re 15 rati b THIN 33 9 7 5135317 3 9 DES I H A E 13 10 DISI 4 14 f 12 TPUX 12 ee eee en 13 hess n 12 ANEL TORT RUE E n 15 n M 16 FGA 4 lr M L August ZU 15 message is located at the bottom of the main analysis window For example if a project has one positive control file that is in agreement with the positive control template the message is PC error 0 1 Example of Positive and Negative samples in concordance with controls If the positive control samples are not in agreement with the template the message will be PC error 1 1 and red lines will indicate the peaks in error Example of Positive sample not in concordance with positive control template Chapter 9 Additional Tools Negative Control Concordance PowerPlex_16 nc fsa Negative Control 491 8 133 x 1 Use View Preferences Forensic to select the file name recognition lt n TTT emnes TE ETT meer mene 2 GeneMarker automatically changes font of negative control file name to red in Filename Tree
36. Exports the currently selected Size Standard in the Size Standard List as an XML file to a specified directory on a local or network computer Exit Closes the Size Standard Editor tool Be sure to save changes to the Size Standard before exiting BestMatch Match Selected When selected the Data Process box appears Each sample in the dataset is compared to the currently highlighted Size Standard in the Size Standard List The green triangle indicators are adjusted to give the best match possible Match AII When selected the Data Process box appears All samples in the dataset are compared to each Size Standard The Match Scores for each sample are averaged together The Size Standard with the highest average Match Score for the dataset is chosen as the Best Match Help The Help menu contains a link to Hot Keys in Size Template Editor Click Hot Keys and the Size Editor Action Help box appears Toolbar Icons Size Template Editor Found in the Run Wizard Template Selection box or in the Tools menu Create New Size Standard Opens the Input Dialog box with a field to enter a new Size Standard name Allows for the creation of a new Size Standard Save Changes Saves modifications made to the Size Standard to the SoftGenetics GeneMarker HID Size Standard directory Delete Deletes the Size Standard that is currently highlighted in the Size Standard List NOTE This action is irreversible Show Dye Allows the user to select a
37. Kinship Analysis using Identity by Descent Data Base Search Finding Nearest Relatives Automated Pedigree Diagrams Paternity Index using 92 August 2013 Chapter 8 Relationship Testing Pedigree Analysis Overview Relationship testing involves testing known genotypes in a familial relationship for simple Mendelian Inheritance laws Since the core loci selected for forensic genotyping demonstrate the Mendelian characteristics of segregation and independent assortment the Pedigree tool in GeneMarker HID is an excellent choice to determine genetic relationships between family members The Pedigree module is designed to aid identification of inheritance patterns and abnormalities for medical forensics All individuals in the Pedigree Tree are directly linked to their corresponding Electropherograms To display the Electropherogram for a sample simply double click on the sample s node in the Pedigree Tree and the Electropherogram will appear in the Charts tab The link between the Pedigree Tree and Electropherogram Charts makes relationship analysis quick and efficient To activate the Pedigree function select Applications gt Pedigree from the Main Analysis window menu bar NOTE The dataset must be sized and a Panel applied prior to using the Pedigree function Pedigree Application Electropherogram Charts Pedigree Tree Samples List 273878 10_06 tes 279870 30 0B tea 279878 20 4 tea 279878 3X 7 ha 27387920
38. PI p1 p2 pi Probability of Exclusion PE PE 1 pl p2 pi PI CPI PE CPE examples PI for D3s1358 0 1404 0 2463 0 2315 0 2118 2 0 6889 or 69 of population are included based on this one marker PE for D3s1358 1 0 1404 0 2463 0 2315 0 2118 0 3111 or 31 of the population are exclude based on this one marker CPI 1 21E 09 or 0 000000121 6 When multiplied the PIs of each marker provide the cumulative PI multiply by 100 to have the percent of the population that Combined Probability of Inclusion CPI Pll x PD x x PE Combined Probability of Exclusion CPE CPE 1 1 PE1 x 1 PE2 x x 1 PE can be included as potential contributors to the mixture CPE or NRME 1 1 21E 09 or 99 9999998 of the population can be excluded as potential contributors to this mixture CPE 1 1 PE1 x 1 PE2 x x 1 PE 88 August 2013 Chapter 7 Mixture Analysis Likelihood Ratios Hypothesis testing and calculations Likelihood Ratios are obtained by comparing the null hypothesis which has a probability of 1 to an alternate hypothesis When there are two contributors to a mixture there are several alternate hypotheses that the analyst may want to test Several scenarios comparing Probability of the Null Hypothesis to the Probability of the Alternate Hypothesis are listed below with sample calculations Scenario 1 The project contains a mixture pr
39. Remove All Removes all sample files from the Data File List field Add Folder Set Channels for Data importing RN Click Add Folder to upload raw data files from a specific folder in Eas the file directory tree Click the Default hyperlink to choose a folder to which GeneMarker HID will always open when the Add or Add Folder buttons are clicked Channels Opens the Set Channels dialog with 4 5 and 6 color tab options and allows the user to choose from ABI MegaBACE and Beckman Coulter standard dye color orders The user can also manually enter BI Grange uz dye color and name The default channel color setup is ABI Set the o Erne fo dye color channels before clicking OK in the Open Data Files dialog box Auto Set Channels Cancel Raw Data Analysis Once the raw data files are uploaded the Raw Data Main Analysis window appears Double click the samples in the Sample Tree to open the individual Raw Data Traces The Synthetic Gel Image displays the unprocessed data in a traditional gel format with larger fragments located on the right The Electropherograms display fluorescent signal intensities as a single line trace for each dye color The signal intensities recorded in Relative Fluorescent 11 August 2013 Chapter 2 General Procedure Units RFUs are plotted along a frame scale in the Raw Data Analysis window with fragment mobility from right to left The largest size fragments a
40. ZOOM IN ZooM IN OuT ZOOM OUT 35 99 103 108 138 August 2013
41. 114 279979 2 08 tea 2798733011 dea 279875 3X C tea 27987910 Y2 isa Ere te 10 5 154 Ladder 1fza Ladder 2 fra Ss Aim 4 qu ati umd Jm 11 Person ID 3 Pies cats M re Liri Sapia Fle ZO Pedigree Tree The Pedigree Tree displays in a standard format the relationship between different nodes or individuals in the sample dataset Older generations appear at the top of the Pedigree Tree younger generations near the bottom Males are represented as square nodes females as circle nodes Horizontal lines indicate mates vertical lines indicate offspring Once a node is added to the Pedigree Tree right click the node to see additional options Select Deselect Individual Node To view the electropherogram for an individual right click their node and select Select Node OR double click the node and the electropherogram trace will appear in the Charts tab on the right To hide the sample s electropherogram right click the node and select Deselect Node When a node is selected in the Pedigree Tree the corresponding sample in the Sample List becomes highlighted 93 August 2013 Chapter 8 Relationship Testing Add Edit Delete Individual If an individual is the first to be added to the Pedigree Tree a family must be designated for the person If the person is already added to the Pedigree Tree right click and select Edit Node to change that person s characteristics To delete an individual from t
42. 7 Click Next in the Destination Location window to install GeneMarker HID in the default folder Click the Browse button to choose a different installation directory ine P3 NOTE The default Destination Location for the GeneMarker HID program is C ProgramFiles SoftGenetics GeneMarker HID version number pee CR eae 8 Click Next in the Select Program Manager Group window to accept the ita Loman Server Manage C inma Gene uk ev and License Server Manager default Program Manager Group NOTE Changing the Program Manager Group default may affect program operability It is recommended to accept the default Imponant The korne Server es produci in a Network Corfuga hon The Licance Server Manage mud ba natalled on the Server ra no to ba 9 Click Next in the Start Installation window to install GeneMarker HID dli win 10 Click Finish in the Installation Complete window 11 The Installation Wizard will close ce 12 Eject the SoftGenetics CD 13 Launch GeneMarker HID by double clicking the GeneMarker HID ia desktop OR open the on menu and de p r GeneMarker HID the version that was just installed GeneMarker program 14 The Configure window will appear Click Run Validation to launch No penc fae codicc has the software 15 If the Run Validation button is grayed out this indicates the 35 day 9 trial period has expired 1 Dota coca 5 August 2013 Chapter 1 Install
43. AB PP 26 Pa Pg AB AC 0 5 P Pe 26 pA Pr AB CD 2PoPcePp AA BB Pops 101 August 2013 AA ALA p 2 pp p 2 pp p 2 From C C Li and Sachs 1554 Chapter 8 Relationship Testing Procedure Although allele calls can be edited in the Relationship Testing tool it is easier to begin a relationship test analysis with good clean traces In order to begin with the best sample traces complete size calling Panel alignment and allele editing in the Main Analysis see chapter 2 General Procedure window prior to launching the Relationship Testing tool File types accepted or generated by GeneMarker HID Pedigree module Pre Ped files SGF TXT BMP The Save to DataBase function allows easy updates of the relationship testing database The default database includes 13 CODIS markers and should be used with complete profile samples Additional markers can be added to the database To launch the Kinship Analysis function select Applications Relationship Testing from the menu bar of the Main Analysis window L M ee 1 Open data file or previously saved Maj Inpro project kake PET Ei Pa iiem Pot Sa LAH Stn FI Carn LF 2 Run Wizard to call alleles ay Img 3 Select Relationship Testing from the wert in r Jr i LA 4 M3 L Lu Applications dro
44. AMEL and Y markers are not included in LR calculations 2 Alleles called in the mixture 3 Probability of Inclusion PI and Probability of Exclusion PE for any selected mixture sample two contributors or three or more contributors Genotypes of contributors Likelihood ratio for selected contributors Combined Probability of Inclusion CPI Combined Probability of Exclusion CPE ug SU aes The analysis should record the hypothesis being tested and any other notes on the analysis in the comments section When the result table is copy pasted or exported as txt files the comments are automatically saved with the report table providing more efficient accountability and presentation of results What to Expect Mixture Analysis and Database Search With Reference File s The File Name Tree indicates that two single a ar CORE source files are potential contributors to the t pepes evidence mixture The analyst may select a EH s n X o ES potential contributor file from the drop down menu at the bottom of the report The genotypes combinations and ratio results are displayed in the result table The Report table at the right of the screen provides the LR for each marker The Combined LR is displayed at the bottom of the screen The combined LR is the likelihood that this individual contributed to the mixture versus a random m person from the population contributed to the jJ I mixture Us
45. All Colors EE Click the Browse by All Colors icon in the Main Analysis aa 3e Eas window toolbar L 179 U21511 D7S220 CSF1 O 158 200 25 300 350 Navigation and peak editing options in the All Color Browser is similar to the Main Analysis window To scroll through samples in the All Color Browser click the drop down menu in the upper right corner and select a sample from the list Once a sample is selected in the drop down menu you can use the Up Down Arrow keys to scroll through samples Icons and Functions Zoom In Out e e Use these icons to increase decrease the zoom aspect of the electropherograms Show Hide Mouse Cross Lines Ft When selected x and y axis grid lines will appear at the tip of the mouse cursor along with the basepair size and RFU value of the mouse cursor position Show Hide Bin Ranges When selected the Bin brackets at the top and bottom of the electropherogram trace will appear Auto Scale Markers When selected the RFU intensities of low peaks are adjusted to match the intensity of the highest peak in the dye color When low peaks are increased the intensity magnification factor is noted in the Marker 2X 8X Print Opens the Print Report settings box See Chapter 6 Reports and Printing Profile Comparison View Applications Profile Comparison View The Profile Comparison View was developed as an ea
46. Allele Editor box appears Adjust parameters and click OK See Create Bin section above for explanation of Allele Editor options To move a bin hold down SHIFT key and left click and drag the vertical grey bar in the center of the Bin to the right or left Let go of the SHIFT key and mouse button and the Bin will remain in place To edit the range of a Bin in the Overlay View click the Trace Mode icon to view the Gel Image Hold down SHIFT and mouse over the vertical white line of the Bin edge When a double headed arrow appears hold down left click and drag the Bin edge to adjust the range Delete Bin Right click the vertical grey bar in the center of the Bin in the Overlay Trace Select Delete Allele The Bin will be deleted from the Panel To delete multiple Bins hold down CTRL key and left click and drag across peaks in the Overlay View A light blue hashed box will appear Right click in the hashed box and select Delete Alleles The Bins highlighted by the hashed box will be removed from the Panel Panel Table The Panel Table displays Marker and Bin information for the dye color displayed in the Overlay Trace frame All columns except Dye and Marker can be edited in the Panel Table Right click a highlighted cell and select Set Value to Column to make all values in the column equal to the value in the highlighted cell Dye Indicates the dye color of the Bin Marker Indicates which Marker the Bin is contained in Size Indicates t
47. August 2013 Chapter 5 Panel Editor the third screen of the run wizard as part of the analysis parameter template in Chapter 3 Align all of the bins within a marker 1 Hold down the shift key 2 Atthe same time place the mouse over the gray marker name bar at the top of the electropherogram 3 The marker rectangle will be outlined in red and the panel name will be in red font when the adjust feature is active 4 Drag the marker to align the bins with the peaks of the allelic ladder 5 Save the panel with signal information the turquoise save icon to enable the major panel adjust feature to work in future projects Align an individual bin 1 Select the gray vertical bar of the bin with the mouse the bar will turn blue 2 Hold down the shift key and click on the gray vertical bar for the bin 3 The vertical bar will be outlined in red and the panel name will be in red font 4 Use the mouse to drag the gray vertical bar to the center of the peak 5 Save the aligned panel with signal information IT Tooly ar d LO HLP D s M MLP D MS n n Pon PO FO Samples 8 5 e o 5 50 a 1 af 5 Pum Fus Options Functions and Icons The following are explanations of menu and icon options in Panel Editor Menu Options The Panel Editor contains three menu options File Tools and Help The File
48. Fa Bs 4 PAT 2 S C S PM S 7 10 PAT 3 C foa PO RO BRAIN r me d B 5 PAT 3M ha Bw 7 PAT 4 Cien Dx i x Bn u PAT 4 j mip n FAT 4 Bis n PAT 5 Cte Bis 14 PAT_S_F fan Bu FAI 5 Misa Bis 1 PAT 6 Cita Bi 1 PAT F isa Bw a PAT_6_M bea Bi o PAT 7 Cte B 15 PAT_7_F fan Ba PAI Laddes 1 14 22 PAT Ladder 2fte Person ID M Icons and Functions New Pedigree File Select New Pedigree File to create a new pedigree with multiple families OR select New Family to add a family to the pedigree file Enter the first family member s information into the New Family New Individual box and click OK to create a new Pedigree Tree Open Pedigree File Launches the Load Pedigree File box Select a PED or PRE file to upload the SMP file will automatically upload and click OK Save Pedigree File Launches the Save Pedigree File box Enter filename and change directory to save the Pedigree Files PRE SMP DAT Show Individual Name When selected the individual ID will be displayed in the nodes of the Pedigree Tree Update Sample Data Select to refresh the Mendelian inheritance calculation after a node or allele is edited Pedigree Parameters Launches the Loci Description box Enter the Affection Locus Description Gene Frequencies Select Markers Number of Liability Classes Penetrances Recombination Values and view the Allele Label and Frequencies ae ce Show Co
49. File field A Windows Explorer omeci Paul Roopa Ris window will appear 4 Navigate to the location of the Panels txt file and click Open 5 Next click the access button next to the Bins Load from File field and locate rcc the Bins txt file 6 Click Open NOTE Select Bins Auto Build if a Bins txt file does not exist 7 Click OK in the Import Panels from GeneMapper box 8 All Panels in the Panels txt file will be uploaded into GeneMarker HID 9 Selecta newly uploaded Panel from the Panel List 10 Edit the Markers and Bins so that they align with the peaks in the dataset 11 Save the edited Panel and close Panel Editor 12 Click the Run Project icon in the Main Analysis window 13 Select the Panel from the Panel field in the Run Wizard Template Selection box 14 Proceed through Run Wizard and data analysis See Chapter 2 General Procedure and Chapter 3 Main Analysis Overview Custom Panel Creation Follow the steps below to create a new Panel based on the dataset currently uploaded to GeneMarker HID Automatic Panel Creation 1 In Panel Editor select File Create New Panel from the menu bar or click the Create New Panel icon 2 The Create New Panel box appears 3 Enter a name for the Panel in the Name field This will be the Panel name that is displayed in the Panel List 4 The Type will by default display the Analysis Type chosen initially in Run Wizard Template Selection The only option
50. Fo Go Pena endo 21 137 SIZE CALIBRATION 445a va va vu ui 0a VA VA EA EU N VON EVEN VA EYE iiaii 35 47 SIZE CALIBRATION CHARTS ccccccecscscscccccccscsccccscscscscscscsccecs 44 SIZE CAL 17 SIZECALLIVIEFHODS 17 SIZE EDITOR ACTION HELP eee ee ee ee eene eene nene eene enun 43 SIZE MATCH viata ava Va n uo Rau Ku o GN n i al n lE eR 43 SIZESTANDARD 16 SIZE STANDARD 74 SIZE STANDARD LIST eee va Ea 39 SIZE STANDARD STATISTICS cccccecscscscscccccscsccccscscscscscscsccecs 45 5 16 33 39 43 12 13 16 SMP FORMAT 96 SOFTGENETICS WEBSITE seien piu uus eal Vu na aun va yoga go 34 SOFTWARE VERSION coses rix dein un ko uu kn na dan capu d uuu 34 SP SATURATED PULL UP 4 eee eee nennen 26 SPIKE REMOVAL 12 14 17 SR SATURATED REPAIRED eee eee eee 26 STANDARD COLOR 455949 VEN RALU V UMEN iiaa 16 START UP SETTINGS cusan esaw 29 START YOUR PROJECT 555i sand Rav ya eR s erPeid eus 222 34 Kp A Ap a EAEAN 25 STUTTER d CER 54 STUTTER FILTER vise vic nta nto stad usto us ku aM
51. H Send the email to tech_support softgenetics com o I The Register ID will be sent to you via email within one business day eee 9 070 J Copy and paste the Registration ID from the e mail into the Register ID Rega field of the Offline Registration tab K Click Register Install GeneMarker HID Software on the Client Computer 1 Proceed with installing GeneMarker HID software on the client x computer as described in the Local licensing Option Installation section above until the Configure Registration window appears Configure Network Client Choose Network Contiquratian xj 2 Click Configure Network Client to configure the client software No piewans eter cont Betore configuring the client computer be sure Matha appropiate service to contact License Server Manager 5 i ab iiy uii e E 3 Click Configure Connection to License Server Manager fromthe i IPS tl Choose Network Configuration dialog box et B Input Server Name or Server IP Address 5 Click Configure and GeneMarker HID software will automatically open if connection is properly established and a license is Lo Seveon E hoose Network Configuration available Upgrade of License Server Manager nuu Activate the License Server Manager console X Proceed with step 3 of Register License Server Manager for GeneMarker HID Usage section above Upgrade of GeneMarker HID Software on Client Computer Install GeneMarker
52. HID software on the client computer by following the procedure in the Install GeneMarker HID software on the client computer section above If the network configuration has not changed the software should activate without configuring the IP address of License Server Questions If you have any questions during installation setup or program operation please contact us at 814 237 9340 OR 888 791 1270 OR email us at tech_support softgenetics com 9 August 2013 Chapter 2 General Procedure Chapter 2 General Procedure Chapter 2 General Procedure Import Data Files Raw Data Analysis Process Data Adjust Analysis Parameters 10 August 2013 Chapter 2 General Procedure Import Data Files After installing GeneMarker HID software you are ready to begin fragment analysis First raw data files must be uploaded to the program Below is the list of file types supported by GeneMarker HID Note many AB 3500 instruments have an option to export hid files with normalized peak heights For greatest accuracy of data analysis GeneMarker HID reads the non normalized peak heights of files that are compatible with the software ABI fsa abi ab1 hid Spectrumedix smd MegaBACE rsd Generic scf sg1 Beckman Coulter esd Procedure GRE EA 1 Launch GeneMarker HID rer 1 C SoftG enetics Forensics Frag Data WISTNPPTENPP16 LADDER fs 2 Click Open Data CS Fiag DataWNISTAPPT B PPIE_ MixD5casel evidence
53. IMB Heterozygote Imbalance Peak intensity does not exceed the minimum percentage of the major peak within the marker as set in the Panel Editor Edit Marker box IHE Inconclusive Heterozygous Peak intensity is within the heterozygous inconclusive range set for this locus in the Panel Editor Edit Panel box IHO Inconclusive Homozygous Peak intensity is within the homozygous inconclusive range set for this locus in the Panel Editor Edit Panel box Save Peak Table Click the Save Peak Table icon in the main toolbar to export the Peak Table information currently being displayed in Excel xls or tab delimited Text txt format All samples peak information for only the dye colors selected will be exported in the table Additionally the user can right click in the Peak Table and select Copy Table Hot Key CTRL C to place the current table information onto the Windows clipboard The information can then be pasted into most common spreadsheet or word processing programs including Microsoft Excel Editing Peaks Double click the vertical grey peak center bar to select a peak Right click anywhere in the Electropherogram or Peak Table to see additional menu options Insert Allele Right click at the place in the electropherogram where you would like to add an allele and select Insert Allele The basepair size or bin name will be applied in the Allele Label and the peak specifications will be calculated and displayed in the Pea
54. Lm frg DOS dua e frag 4 Fes haa DOT Ts freg_006_O01 Fea ag OU Egi ita hag 008 a01 Fea frag DOS fey Uil GO Ies E ha 1 Fi l ra 015 ET fra 8 imag 013 tos B iaa 014 onii m Cancel Size Standard List The Size Standard List contains all pre defined Size Standards and any custom made Size Standards Single left click a Size Standard in the list to select it The Expected Size Standard trace and Size Table will appear on the right Additional Options To see additional options for each Size Standard right click the Size Standard name and the right click menu will appear with the following options 39 August 2013 Chapter 4 Fragment Sizing Standards Delete Size Standard Select Delete to delete the Size Standard from the Size Standard List and from the SoftGenetics GeneMarker HID Size Standard directory NOTE This action is irreversible Export Size Standard Opens the Save As window Choose a directory folder and click Save The Size Standard will be copied to the selected directory and will also remain in the Size Standard List and SoftGenetics GeneMarker HID Size Standard directory The Size Standard will be exported as an XML file which can be opened with Internet Explorer Microsoft Excel or Notepad Reload Size Standard Click Reload to undo editing changes to the Size Standard The most recently saved Size Standard will be restored NO
55. PANE Cle GU ON osuere c oio ates ec Pede od te cio rerba vata d b Peedoe ideas date ovv pe vtae bow epa ea VE n ode eade uie pgs 56 Adjusting Panels Manual and Automated Panel Calibration sessi 57 OPTIONS FUNCTIONS AN Ds CONS 50i s os xao rds On S vu ru RR T nc bin do oh a DAS vU lae PU VO ae AU RAO CMS 58 MICU 58 TOO ON oos tube dae tanec Teh s reb t aoa NS 59 Saving apanel with sigbal ia orma ON as e nob E RR bt b e b ER RU eet 60 Project PANE Ec 61 WAATI EXPE T M LM MM EUM LI tx ME EM LE 61 CHAPTER 6 REPORTS AND PRINTING eoi ei eea ia pes eux E De Vena OS Gu EE DE One pa D Pone t uxDe edu xs v Una bp Onde va Ta Dee ucro De PO a De QUEEN SUP QNS 66 REPORT TABLE seit eei e cc IE a c LP So 67 AGI EISE ove se veiba beue tp Lor rM M iU E A oU PUE 67 FOrENSIES ar Hc O tbe 67 SERITUR 68 ROO TIBI ORT t RUNS MM S LR LO S 69 R rc TD 70 PRINT REPOR oderit esa d odeur Ups vtae Dr a Ner radere eua oe 71 REPO COMON OPUN tmr 72 leons HO FUNCIONS oer oai R 73 GOD IS REPORT cg aces seacoast 74 SAVE PROIEG asocio
56. SG1 if exporting a five color trace click Export to SCF if exporting a four color trace Export Electropherogram Tools Export Electropherogram The Export Electropherogram tool allows the user to export the trace images to a specified folder Procedure 1 Use a dropdown menu to specify the output folder 2 Specify the prefix and suffix for the exported file name The full file name will be Prefix Sample name Dye name tSuffix Extension name 3 Select samples Dyes and Image Size 4 Use a dropdown menu to specify the export format JPEG and PNG are both available PNG is recommended 121 August 2013 GG Convert Text to Binary File Maxintencty Recommended Rano 4 65535 AS 46 I Conderan Frame Ga Load Test fie Une Asie Condence Framex Output Trace Graph Fodar IC VU rer V5 on enenies D esb ep Flelatonshep Teva DC E yenay Daa 5 Hebe Salle Select Sample 1 Che Width hug J r ri Hegt 758 inet w PAT 5 C ha h Dus iv PAT F ta Epon Foma NGA l Chapter 10 User Management Replicate Comparison Tool Tools Replicate Comparison Tool Many labs choose to run and process multiple replicates of their samples This ensures that a genotype is still available in cases of contamination allele drop out or reaction failure Concordance between replicates can then be used to deduce the genotype of the sample The Replica
57. Test TestPed pre l Ns d Jm T A ER AP Fawip i22 2 Famip2 Jones Marker 3 D75820 Search d amp EE L net Samples Charts k Ei dI 4 P 258 3 1748 x 300 a ERR m Ed 2E a 27 9875 10_06 f a 2646 5767 X 245 z 255 260 25 270 2800 205 290 295 300 278878 30 11 58 245 250 Person Mame Jon Sample File 279879 20 4 58 100 August 2013 Chapter 8 Relationship Testing Kinship Analysis using Identity by Descent Overview STR profiles of two individuals can be compared to determine the likelihood that they have a specific relationship versus the likelihood that they are unrelated Kinship analysis compares STR profiles from individuals to determine likelihood of a family relationship versus the likelihood that two individuals with these STR profiles are unrelated The formulas used to calculate the level of kinship depend on 1 Probabilities that 2 1 or 0 alleles will be shared IBD identity by descent given a specific relationship 2 The probability of a specific genotype X given genotype Y at all loci under the conditions that X and Y have 2 1 or O alleles IBD GeneMarker HID uses established rigorous statistical analysis Kinship formulas from Brenner 2004 Eisenberg and Planz 2007 to calculate probabilities and likelihood ratios for different relationship levels including Pare
58. The data is sized peaks are filtered and the Panel is applied Mere 12 Click OK when the Data Processing box is finished i VALE m Functions Ha k parcel Allelic Ladder Permits the selection of a sample containing an allelic ladder If the user selects one ladder the ladder will be in bold font and is set to the top electropherogram in the Main Analysis window All samples will be analyzed using this selected ladder Allele Evaluation Peak Score User definable confidence level of the allele call Peak score is an algorithm that takes into account signal to noise ratio and peak morphology Rejected samples appear in red samples that need to be checked appear in yellow and samples that have passed appear in green Auto Select Best Ladder GeneMarker HID identifies ladder samples in the dataset as defined in the View Preference Forensic gt Ladder Identifier field Ladder samples are then compared to the chosen Panel Each ladder that is within the range of the selected panel will pass and appears in bold font in the Sample File Tree Auto Select Best Ladder will analyze each sample file with the passing ladder that best matches that sample The print report provides the file name of reference ladder used for each sample Auto Panel Adjustment 18 August 2013 Chapter 2 General Procedure When selected the Markers and Bins of the chosen Panel will be aligned with the peak positions of the Ladder samples in t
59. at left select Edit Comments to add new comment or select a comment from the list Quality Reasons Indicates the reason why a peak received a Quality rank of Check or Undetermined For explanation of the two and three letter codes see below OR click the Help icon above the Report Table 25 August 2013 Chapter 3 Main Analysis Overview LS Low Score Quality Score is based on Signal to Noise Ratio and Peak Morphology and the Pass Check Reject ranges are set in the Run Wizard Additional Settings box OL Off Ladder Peak is outside of the marker range OB Out of Bin Peak is within the marker range but outside of a bin BC Bin Conflict More than one called peak present within a bin SR Saturated Repaired Intense peaks with characteristic morphology are identified and repaired for allele calling SP Saturated Pull up Intense peaks may cause pull up or additional peaks to appear in other dye colors PL Beyond Ploidy When the number of peaks identified within a marker exceeds the maximum number of peaks expected as set in the Panel Editor Edit Panel box LO Low Intensity Single peak called below the Minimum Homozygote Intensity threshold because a second peak was detected above N x percentage value as set in the Panel Editor Edit Marker box HI High Intensity Peak intensity approaches and or exceeds the maximum peak intensity filter as set in the Run Wizard Data Process box
60. be lost when data is re analyzed Re analyze with Run Wizard To re analyze with the Run Wizard tool simply click the Run Proj ect icon in the Use Old Calibration Being closed in 23 seconds X main toolbar The Run Wizard will launch and the most recently selected A Size Calibration parameters will be displayed Adjust parameters as necessary and click OK in The intemal lane sizes have been calibrated in the samples You may check Call Size Again to recall the sizes the Run Wizard Additional Settings box The Use Old Calibration box will appear as with the option to Call Size Again Only select Call Size Again if the Run Whattodo Wizard Template Selection Size Standard selection was changed or any of the Run Call size again Wizard Data Process Raw Data Analysis parameters were changed Click the Apply to All button The Data Processing box will appear again and the data will be re analyzed with the new parameters Re analyze with Auto Run To re analyze with Auto Run first select Project Options The Project Options Settings box will appear This box offers all the same parameters settings as are available in the Run Wizard Use the tabs to view the Template Selection Data Process and Additional Settings boxes Click OK when finished Next select Project Auto Run The data will be re analyzed with the new parameters NOTE The Additional Settings Allele Evaluation Peak Score parameters can be changed in the Project Opti
61. colors This feature includes a 2 Dimensional and a 3 Dimensional view of the selected samples Relationship Testing Contains tools for familial search identities duplicate samples and potential near relatives from the relationship testing database provides likelihood ratios for each match kinship analysis parent child sibs half sibs aunt uncle grandparent and cousin and automated pedigree drawing with deduced genotype of missing parent based on child ren and available parent where possible Cell Line Verification This Application is currently under development It will function in a similar way to the Relationship Testing and will provide immediate authentication of cell line based on published genotypes Mixture Analysis GeneMarker HID identifies the presence of potential mixture samples designates allele peaks and calculates peak area or height ratios in the main analysis screen The Mixture Analysis Application is activated from the Applications menu in the main analysis screen Mixture analysis identifies the 32 August 2013 Chapter 3 Main Analysis Overview mixture samples and any single source contributor samples in a file name tree considers all possible allele combinations calculates the Mixture Ratio residual score heterozygous imbalance for each genotype combination and calculates the likelihood ratio for single source samples that are potential contributors to the mixture searches database for single source file
62. deleted by right clicking the username and selecting Delete User NOTE Only the Administrator can add and delete users Access Rights of User Types My Password Access Rights User Type Launches the Change Password box where the user that is logged in Y Edt lees iw Inse eles iv Del llel Lab M can enter a new password The new password must be entered ndo Mn retia a ivi Confir twice to ensure accuracy V Unconfrm Alleles Mw prie iHa v Recall Alleles Access Rights Re Run iv Set Run Option Users Launches the Access Rights of User Types box where the different peer access rights available to each user type can be identified Clicking V Comment Samples the Set Default button will return the Access Rights for the User comment project v Save Project Type selected back to factory defaults FERE NOTE Only the Administrator can change Access Rights for a User Update Sofuare e Type re alysis Parameters Change User Set Default Cancel Prompts for a confirmation of action then launches the Login box 129 August 2013 Chapter 10 User Management History The User Manager History tab monitors user activity associated ipi x with the user manager function User Manager History Settings DateTime User Events Comments Date T Ime 7 20 2011 1 28 13 PM Admin Add new user tech 1 jones Records the computer s date and time for the activity n 7 19
63. ee eene eene eee sepan 18 92 AUTOMATIC PANEL CREATION nha Ra ka ER REA FRU 56 AUTOMATICALLY 5 31 AUTOMATICALLY SCROLL CHARTS TO ALLELES WHEN SELECTED IN ize gl 31 B BASELINE SUBTRACTION eee ee ee ee ee eee nennen 12 13 16 17 BC BIN OCC P 26 133 BECKMAN COULTER 11 BESTIVIATCH aiisssdst asia p Eo po E Ead M AERA SEL ERE REAPER AERA 41 BESTMATCH MATCH FALL von 43 BEsTMATCH 43 BIN BOUNDARY icit etre xke Ex MEE EFE CrkEE EK Ek ERES 54 pe cg cm ciio triet 54 BIN RANGE scsesccapesas 53 68 BINNING conso tum tat akt 53 54 io 63 35 99 104 108 BROWSE BY ALL COLORS iiriee seva seas ea rarae rara sais agre ser eise 36 C CALIBRATION PLOTS s c sem PLE NM 46 4 PE TII LETT 35 CALL ALLELE ICON casas satin aor Ek rv REEF RM REA 19 CHANGE PASSWORD zi ev ore aure rea au 129 CHANGE USER Mmmm iainih 129 CHART
64. green sheet those with a low Match Score have a yellow sheet Samples where size calling failed receive a red strike through To analyze how each individual sample was matched to the Size Standard selected access the Size Calibration Charts Within Size Calibration Charts the user can modify how each sample was sized and view the statistical information for disabled Size Standard peaks Sample List The Sample List includes filename Match Score and disabled peak information for each sample in the dataset Sort the list by single left clicking the column header The list will re sort in ascending or descending order based on the values in the column selected Single left click a sample to view its Sample ILS and Calibration Plots on the right OR use the Up Down Arrow keys Right click the sample row and select Mark as Failed to disable the sample select Unmark Failed to reverse the action Disabled samples will appear greyed out in the Sample List Score The Score column displays the sample s Match Score which corresponds to the degree of pattern match between the sample s ILS and the Size Standard selected Perfect matches receive a score of 100 no correlation receives a score of 0 and the sample is considered to have failed size calling Disabled Size Columns The Sizes that were disabled in the Size Standard see Size Template Editor section above will appear as column headers in the Sample List If no Sizes were disabled then only the Sam
65. hgust 2013 E d ooo ooo n 2 31 Nat hd haa sE RI Gell 21 Chapter 5 Panel Editor Project Panel VM Once a panel has been aligned and saved with ladder signal information 0 a a x it can be set as the project panel This panel has signal information and is used to automatically adjust to multiple ladder files in a project Right click on the panel name in the list of panels and select Set As Project Panel The panel name will be displayed at the top of the panel list under the heading Project Panel Exit from the panel editor and the project panel will be applied to the project Samper a 2002 09 26 A01 LADD 2006 03 26 Am LADD 2008 09 26 LADD 2008 0926 A01 LADD 2008 03 26 LADO What to Expect Once a Panel has been created aligned and saved with signal information it can be applied to the dataset Save the edited Template Panel with signal information in Panel Editor then exit the Panel Editor If the Panel Editor was accessed via the Run Wizard Template Selection box icon then the selected Panel will appear in the Panel field If the Panel Editor was accessed via the Tools menu then click the Run Process icon in the Main Analysis toolbar The Run Wizard will appear Select the Panel from the Panel drop down menu in the Run Wizard Template Selection box Proceed through the other Run Wizard boxes and click OK when the Data Process window is complete The Panel will be applied
66. in GeneMarker HID is HID Analysis Type 5 Select Automatically Create 56 August 2013 Create New Panel _ 3s Name TestPanel Parameters aa i zi Iw Fixed Bin width 0 5 Method Manually Create Automatically Create f Use All Samples Use Selected Samples Cancel Chapter 5 Panel Editor a Use All Samples will create a Panel based on an overlay of all the sample peaks in the dataset b Use Selected Samples will create a Panel based only on the samples selected in the Panel Editor Sample List Click the double arrow button to expand the dialog box and see additional parameters If required check the Fixed Bin Width option and enter a value for the left and right Bin ranges When finished click OK The new Panel will be created and added to the Panel List NOTE New Panels are created based on the Max amp Average View Mode More intense peaks are given higher priority for Bin placement when peaks do not overlap perfectly 10 Edit the Markers and Bins as described in the previous section Panel Editor Overview 11 Follow steps 10 14 above VOND Manual Panel Creation The Panel name will appear in the Panel List however no Markers or Bins will be associated with the Panel Follow the steps in the previous section Panel Editor Overview to create Markers and Bins 1 In Panel Editor select File Create New Panel from the menu bar or click the Create New Panel icon 2
67. in the Main Analysis window if Project Apply Sample Grouping was chosen 10 To navigate by group in the Sample File Tree hold down the CTRL key and hit the PageUp Down keys Sample groups will be opened consecutively Icons and Functions je Load Files Opens a directory window where raw data files can be located and uploaded to the Filename Group Editor Add Files Opens a directory window where additional raw data files can be uploaded into the Filename List field Remove Files Removes any files selected in the Filename List Select multiple files to remove by holding down the SHIFT key and selecting additional samples 118 August 2013 Chapter 9 Additional Tools Save Groups to File Saves the filenames of the samples paired in the Matched Groups field Samples identified as Controls will be in the first column of the Matched Groups tab delimited Text file Match by Sections Automatically separates the sample filenames into groups based on the specified Section Separators Group Identification Identifies how to match the filenames into groups based on the section entered into the Compare by Section field The section of the filename specified will be highlighted red in the File Name List Control Identification Identifies which section of the filename contains the reference vs sample information based on the section number entered in the Match to Identifier by Section field The section of the filename specified w
68. include dye colors data type and the directory to output the trace files See Chapter 9 Additional Tools Export Electropherogram Allows the user to export the trace images to a specified folder Magic Wizard Contains three option boxes Start Your Project Run and Report Start your project Start Your Project Hd Allows the user to easily access the Open Data or Open Project G Upen Data upload windows The user can also re open the four previously Open Project opened projects by selecting the black arrow next to Open Project Run Selecting Run launches the Run Wizard Selecting AutoRun will process the data automatically with the process options currently selected See Chapter 2 General Procedure Report Allows the user to Save Project or Print Report Selecting Print Report will launch the Print Report Settings box See Chapter 6 Reports and Printing Show Last Event Opens the last active Data Process action Help Menu dii Help Launches a searchable version of this manual User Management About User Management Allows an administrator to assign access rights to different users Also used to set up the password protection feature See Chapter 10 User Management About Displays information specific to the version of GeneMarker HID running on the computer Also contains links to email Technical Support and the SoftGenetics website Main Toolbar Icons Open Data Opens data input dialog box to be
69. is 11 12 13 and the intensities are 1000 1000 and 200 one of the valid combinations could be 11 12 and 13 Q In this case the Major Mx is 1000 1000 1000 1000 200 100 and the minor HIM is 100 200 or 0 5 But if the intensity of 13 was less than 100 the intensity of the drop out peak would be set at the same intensity as 13 Contributor detection threshold allows for possible contributor when there is an incomplete profile Preloaded Allele frequency values are the same as those described in Chapter 8 Relationship Testing The edit icon allows individuals with access right approval to easy add or remove other population allele frequency txt tab delimited tables see chapter 8 for file format template Allele Freuqency Tables Allele frequency tables for major US populations may be selected from the drop down menus in the Select Allele Frequency Settings box If results of all populations are preferred select the Use all Populations box and the final report will append with the results using each of the tables sequentially The Delete button may be used by individuals with access rights to remove any population frequency tables that are not needed by the laboratory Use the Open folder icon to import formatted allele frequency tables for other populations Skip Unmatched Markers in Cumulative LR calculation allows for LR calculation in the cases of incomplete profiles Overlay when Show Invalid Combinations is selected all po
70. nul e veau eut 36 LINKED USER 34 LINUX SPINE DO DU T 5 LO LOW INTENSITY uisi oxke aut paa rh xPP PEE prb FE ree 26 LOCAL SOUTHERN 4 5eusshs uuut uk vk E EEEE dE kd 17 Loci DESCRIPTION FILE cccccccccecscscscscscccsccccecscscscscscscsccecs 96 LOGIN m M 129 LOW TEMPLATE FLAGGING aPu EFE SEEN ERE es ERES EF 53 ES LOW SCORE eub ea 26 M IMACINTOSH visa iE SERRE RARO EDU VIRA du 5 MAGIC WIZARD VR V Fav eau rEY UE 0s 34 35 60 IVIAIOR VIX Voas iE Fax RP Va ER FE EP ES REN 87 MANUAL CALIBRATION 656553 eaa y ava asa Ee Sua Few Ea Sao esa revo 47 MANUAL PANEL 57 12 MANUAL SELECTION OF 18 5 29 53 MARKER DROP DOWN 35 IVIARKER OPTIONS i515 vens aYa T Esau Ev
71. o an nu Pu aiu e a ud ko Kan Ru nu 52 EDIT SIZE DS iua dua ouo AUOD IO OT 40 EDITING PEAKS cosi ioo 26 28 EDITING SIZE CALE ieiuna nana tcasstectcdsntastedeviestedessastsasniasscdns 45 ELECTROPHEROGRAM esee eese eee sees eese ee eese eese see sess eese sue 23 ELECTROPHEROGRAM 5 24 ENABLE SAMPLE GROUPING 5 5535 ova yes ER ewe 31 ENHANCED BASELINE 5 17 ENHANCED SMOOTH suut ra regole 16 EVENT siii teneis 35 e 29 EXPORT ABI SIZE STANDARD eee ee ee ee eene nene nennen 42 EXPORT CALIBRATION DATA eee ee ee ee eere eene neenon eene 46 EXPORT GODS 25 iud gata send cauaa aux deua MA a a aid aoa tait 32 74 EXPORT 34 121 EXPORT PANEL 52 59 EXPORT SIZE STANDARD pu anta uva vun vaa Ta uas 40 42 EXPORT THE PROJECT PANEL e eee ee ee eee ee eee eee eee nue 59 134 EXPORT TO SCP X VENT VUE VIR VA EWEAS EN EWEN NE TUER 121 EXPORT TO SQL s vine ne uua oaa 121 EXTEND DIPLOID HOMOZYGOUS eee eee eee eee ee ee eene nenne 67 EXTENDED PEDIGREE FILES sccsvssovessvsvs
72. of the Report Table The Report Bin e mm mm Columns box will appear it dua Ce EA Bi 554 as m 035138 Gwen Display Bins mS Bi as n5 nh 0351358 By default all Bins will be selected with a checkmark at the A pausa beginning of the row Individually deselect Bins for exclusion from menm the Report Table by single left clicking the checkmark box To m s ime oem orm i E 02 m a a deselect all right click any cell in the Report Bin Columns box and select Uncheck All To deselect only a few Bins left click a cell to 5 T ap he mr Dis Tar 05 05 bE highlight the row then hold CTRL or SHIFT key and select additional rows Next right click and select Check or Uncheck to include or exclude the Bins respectively Click OK in the Report Bin Columns box when finished and only the Bins with checkmarks will be displayed in the Report Table Merge Bins To make two or more Bins become one Bin single left click a row to highlight it Next hold down SHIFT key to select additional rows Right click the highlighted rows and select Merge Bins Hot Key CTRL M Click OK in the Report Bin Columns box when finished and the selected Bins will be averaged together Only Bins immediately adjacent to one another may be selected for merging Only the height and area for the first peak in the new merged Bin will be reported Peak Table The Peak Table Report Style dis
73. or copies thereof are not transferred to another party or computer unless all copies of the Update are also transferred to such party or computer and you acknowledge that any obligation SoftGenetics LLC may have to support the previous version of the Software may be ended upon availability of the Update 6 LIMITED WARRANTY SoftGenetics LLC warrants to the person or entity that purchases a license for the Software for use pursuant to the terms of this license that the Software will perform substantially in accordance with the Documentation for the ninety 90 day period following receipt of the Software when used on the recommended hardware configuration Non substantial variations of performance from the Documentation does not establish a warranty right THIS LIMITED WARRANTY DOES NOT APPLY TO UPDATES FONT SOFTWARE CONVERTED INTO OTHER FORMATS PRE RELEASE BETA TRYOUT PRODUCT SAMPLER OR NOT FOR RESALE NFR COPIES OF SOFTWARE To make a warranty claim you must return the Software to the location where you obtained it along with proof of purchase within such ninety 90 day period If the Software does not perform substantially in accordance with the Documentation the entire liability of SoftGenetics LLC and your exclusive remedy shall be limited to either at SoftGenetics LLC option the replacement of the Software or the refund of the license fee you paid for the Software THE LIMITED WARRANTY SET FORTH IN THIS SECTION GIVES YOU SPECIFIC LEGAL R
74. the Match Score calculation The Match Score calculation is updated when Update Calibration is selected NOTE Add Peak is only available when no other green triangle peak indicator is selected Fix Size When selected the Calibration Editor box appears Enter the correct basepair size of the peak and click OK NOTE Only Sizes that occur in the selected Size Standard can be entered in the Calibration Editor Size field The peak will be fixed at the specified basepair position and all green Calibration Editor EX triangle peak indicators to the left and right of the fixed peak will be adjusted to correctly align with the chosen Size Standard Peak 5086 ai The Fix Size feature is useful when the selected Size Standard has uniformly pice spaced peaks and the sample ILS has additional peaks due to pull up or other experimental abnormalities thereby influencing the pattern recognition Cancel algorithm NOTE Fix Size is not active for manually added peaks or peaks outside the Size Standard range Reset Peaks Select Reset Peaks to eliminate manually added peaks and or extra green triangle peak indicators after Fix Size NOTE Deleted peaks will not be recalled when Reset Peaks is selected Update Calibration After editing peaks in the Sample ILS select Update Calibration The Match Score for the sample will be recalculated based on the edited peak indicator p
75. the peak to the right side of the peak or in the allele label in the Electropherogram Gel Image Gray for Single Dye When selected will display and print the gel image with a black background and white bands When deselected the gel image will display a black background and colored bands depending on dye color chosen to view NOTE When all dye colors are selected the bands in the gel image will be displayed in color regardless if this option is selected Background in White Only available when Gray for Single Dye is selected Will invert the gel image so that the background will be white and the band fragments will be black Forensic The Forensic tab allows the user to determine the display of Ladders and Controls in the Report Table and to establish file labeling conventions for Ladders and Controls Show Ladder Samples in Report When selected the samples designated as Ladders by the Ladder Identifier field will appear in the Report Table Show Control Samples in Report When selected the samples designated as Positive and Negative Controls by the Positive Negative Control Identifier fields will appear in the Report Table Mark Deleted Edited Peaks with Symbols when selected samples that are deleted are marked with an y at the top of the peak Samples that were edited are marked with an E at the top of the peak Label Peak Ratio Select from displaying peak ratio from height or area in peak flags on the electropherogram Ladder Iden
76. to the de convolution profile from the mixture Magn rii age ag Doria Meca victim Mna irei 2 Dg Anasa Mike Mac sample Select the file to display the LR results in the Report Table 83 August 2013 Chapter 7 Mixture Analysis dual Mare HIM T Miror HIM Two Coetibuters 1113 1114 087 n358 559 Ne Mane y pH MixfBcasez ev 1113 1414 0 93 Dong 036 113311 3573 36907 423 Tren ACC TE Wana J6XE 3 n5 LX NC UO GS 381011 43189 911 12137 ee Late MA 5 7101213 25300 1213 14 426 ELO LL 0161358 516 arias 1516 x M wm rHO 69310 0425 10 nsu a a EMO s HSI 451214 02 370 amaw B65 EE Rae L LEO 307 os messis 3101132 03338 512 13517 jen rss OUS 90 ans o5 D251310h 16172124 oas de 1524 121258 Me eB oe D 95433 127314 1214 4513 ya Mn Gio A 040 WWE 151519 017263 gen 3 23330 2 TPOX 3310 11 oases g5 BN 3842 S EN au zi aug piesi 12051718 02354 4 1215 ME 131 1414 ag 0 0054 AMEL ay x UR um pases 3111213 079333 amar 1712 3517 a30 qi 03 51 FGA 202324 015 93 gsi 2321 5903 1738 1215 oe nonz 90 0 82 A TE iVE 1139707 1 34 18 es x 1 00178 Cumulanwe TEO S7E 07 Es XN D 0s anm 063 163 osae 83 86 0003 033 166
77. up Removal Automatically removes peaks caused by wavelength bleed through to other wavelengths This option is selected by default in the Run Wizard Manual Pull up Correction This allows the user to manually adjust larger pull up peaks in case the Auto Pull up Removal function has not corrected the problem It is recommend to de select Pull up Correction in the Run Wizard when using this function 2 d Derivative Trace This feature reduces high background noise and sharpens peaks Baseline fluctuation caused from dye blobs or the DNA template in PCR can also be reduced with this function It is recommended to de select Spike Removal in the Run Wizard when this function has been activated 12 August 2013 Chapter 2 General Procedure What to Expect The raw data correction icons can be selected individually in the Raw Data Analysis window The images below demonstrate how the data will look before left image and after right image the parameter is applied Range AutoRange Analyzes from 0 to end of trace for size call Manual Range user defined range Right click in gel image and select Get Start Point Gel image Get Stat Point Automatic ali Slaton inn Smooth Fourier frequency transformation FFT to determine frequency domain Use only top 40 of lowest frequencies Smoothing broadens peaks and therefore you can lose resolution Enhanced Smooth Same as Smooth but use only top 2076 of lowest frequencies Basel
78. 1 FGA 34 2121 n 09 doa 0537 ig I X 233 im anes 047 1 amp 3 FGA Xi Ma o nw ne rea FGA 074 7 28 os RA m E aa AE amp oj r 4 mt had f P iis n Akiti Ti hypotheses eo mainais fv ene nuo oder people conitured I in moni d Orituco Z ME emen Mage Mom gt F onat aym spo Ha Me dm Clumber i PEE pave 1 150007 LI Contesting both contributors to the mixture Icons and Functions Mixture Analysis Parameters Launches the Mixture Analysis Settings Dialog Box Edit Database Navigates to the Edit Database function in Relationship Testing Submit Genotypes to Database Allows direct submission of reference file genotypes to the relationship testing database Locus Name Group Editor Allows the user to group different format names for recognition of a single marker Toggle Toggle between 1 all possible allele combinations black and red font and 2 only the allele combinations that conform to the mixture analysis parameters black font Show Color Allows the user to select all colors to view hide all colors or choose a single dye layer in the trace view Choose a single dye by single left mouse clicking on the icon Marker Marker All Markers Select a Marker or Locus to view in the Electropherogram Charts 86 August 2013 Chapter 7 Mixture Analysis arn Submit to Relationship Tes
79. 15 0 0 Check 406 2 IMB 128 9 5852 1 00 35227 D351358 16 0 1 Pass 500 0 133 0 2318 0 40 13563 D351358 17 0 2 Check 220 2 IMB 1762 5125 0 77 24030 THO1 7 0 0 Pass 500 0 180 4 6626 1 00 40931 THO1 8 0 2 Pass 500 0 2283 7796 1 00 43069 D135317 11 0 1 Pass 500 0 2323 3723 0 48 21043 D135317 12 0 1 Check 500 0 IMB 2723 1518 0 40 10558 D155533 10 0 2 Check 123 4 IMB 276 4 3755 1 00 24684 D16S539 11 0 2 Pass 467 9 280 5 2026 0 54 13283 D165539 12 0 1 Check 187 6 IMB R 316 0 1921 1 00 14134 D251338 17 0 0 Pass 147 9 332 4 832 0 43 5840 D251338 21 0 1 Check 44 0 IMB Page Up Paa m D Modified 6 samples PC error 0 1 NC error 0 1 Ladder error 1 1 Failed 0 Flagged 3 5 1 Ladder and controls are automatically T xm identified based on no enclature Start up Settings Display Settings Forensic Others specified 1 View Preferences Show Ladder Samples in Report Show Control Samples in Report 7 Mark Deleted E dited Peaks with Symbols Forensic E iw Label Peak Ratio 2 Action summary presented at the bottom gt Label Peak Height Ratio Label Peak Area Ratio of the screen indicates number of CRET ADDER samples automated positive and negative Positive Contiol Identifier PC Negative Control Identifier NC control concordance ladder flagging see chapter 5 Panel Editor number of samples failed and flagged caused rule firing Cancel 3 Samples that meet the mixture criteria are identified
80. 2011 8 45 08 AM Admin Log out 7 18 2011 10 07 45 Admin Log in User 7 15 2011 3 01 14 PM Admin Log out apie 7 15 2011 2 00 27 PM Admi Logi Identifies the username of the person that performed the 745 2011 14843 Admin Lo ou 7 15 2011 11 21 57 Admin Log in activity 7 15 2011 10 47 02 Admin Log out 7 15 2011 10 15 07 Admin Log in 7 15 2011 10 15 07 System Add administrator Events Records the user manager activity that was performed Comments Gives additional information for the event that was performed t Run User Protection For example if a user is added then the username of the person that was added is recorded under Comments Print Use the Print button to preview the User Management History print the history or save as a png pdf or jpeg file SoftGenetics User Management History 7 20 2011 1 28 16 PM GeneMarker HID v2 1 2 Page 1 DataTime User Events Comments 77207 2011 1 28 13 PM Admin Add new user tech 1 jones 7 20 2011 11 41 21 AM Admin 7 19 2011 9 04 40 AM Admin 7 19 2011 8 45 09 AM Admin 7 18 2011 10 07 45 AM Admin 7 l5 2011 3 01 14 PM Admin 7 15 2011 2 00 27 PM Admin 7 15 2011 1 49 43 PM Admin 7 l5 2011 11 21 57 AM Admin 7 15 2011 10 47 02 AM Admin 7 15 2011 10 15 07 AM Admin 7 l5 2011 10 15 07 AM System administrator Settings The User Manager Settings tab contains additional options for the User us user Manager admin toaged in SCEA Management func
81. 224 nza 6 owe 037 Z PGA 223 no is 03 033 m w 0M 085 083 Pid X34 na asm SC L m Q 4 p v u er Tre i VOTER AA Searching database fat malth to mejot resulting hom denorvokition cl Micase mvutence Corata Wane piy Average Maan Mix case evidence Aao Mayas Indre X12 346 18 11 137 17 file Ede a Bi mi Files can be saved in CODIS format by using the Export a i a re ee FlesNSoltGeneticiNGeneM aer HID 2 6 PAT 1 Ffsa CODIS icon from the Genetic Database Editor E PATA Mee Lut Contro 5 6 PAT ha PAT 2 PAT 3 C tea FAT 3 Frise PAT 3Mfsa PAT_4_Cica PAT_4 F isa 4 lea 5 Clea FAT 5 Fitee FAT 5 Misa amp 13 PAT 6 Fla PAT 5 M za FAT 7 Clsa FAT 7 PAT Lad 166 ID Range t PC postive contiol fea Ideris LADDER tea MIAD6cace2_ vitm isa XYZ MIXD5cese2 evterce Mie D ng Sto Mor Db Brg Female FOV Isa D Eng Made GO4 tea BRASAS BE Hypothesis Testing Options To calculate likelihood ratios from a comparison of the null hypothesis to alternate hypotheses the analyst should select contributor 1 and contributor 2 from the drop down menus at the bottom of the analysis screen Check the appropriate box es under the Contested heading to contest contributor 1 or contest contributor 2 or contest both c
82. 29 AFFECTED sat vss dena c ves eus ve 94 114 AFFECTION 5 5 98 AGAROSE 39 ALLELE BOUNDARIES i5 ra ik a wa KE ERE 60 ALLELE ERR EE ERA ERE Ada ERR 18 ALLELE CALL FOLDER iss 21 ALLELE EVE VEN 27 70 ALLELE EVALUATION 18 ALLELE LABEL 29 98 ALLELE teu AER EU r EE E rE DU EErEE EE REED E EE 67 ALLELIC LADDER ev RH Lo e E Go Rau RD BU RR 18 eva a o 16 5 54 5 31 AUDIT ge tiusiiist 131 AUTOFIT V GEN 47 AUTO IDENTIFY ED DEEP 31 AUTO PANEL 18 AUTO PULL UP REMOVAL 12 AUTO RANGE 16 18 AUTORUNGS 32 AUTO 5 5 36 72 AUTO SELECT BEST LADDER 4 ee ee ee ee ee
83. 50 250 150 250 350 Q5 250 Dmm gi n alb ladies 8 rad 8 0S ddaa HE adder 2c 6 rec D rc am o7 54 i L ye f me B 7 ae T T L oe 1515 et 156 256 z 2222272 zs Chapter 5 Panel Editor oom inda ac 1 The first two allelic ladders show a migration shift Globalfiler Panel GF ladderl and ladder 2 left When Globalfiler Panel is selected from the list the electropherogram displays the aligned bins for this ladder file After the Panel is applied to the dataset the Markers and Bins appear in the Electropherogram and Report Table In the Electropherogram the Markers are horizontal grey bars the Bins appear as dye colored brackets above and below the trace and the center of the peaks are marked with a vertical grey bar Peaks that fall outside of the Markers are marked Off Ladder OL Peaks that fall within the marker but outside of a bin are marked OB The allele ladder that best matches a sample file is used by the Auto Select Best Ladder in the Run Wizard The file name of the ladder used for a sample is displayed in the allele report See Chapter 6 Reports and Printing EPI P View Project Applicatinns Took Help PPM S FOB BW me ind a I Raw Data GF ladder AT1 01 hid Allie Call BB DOENTE c thid
84. 65 vata va ua n UE UA ULM 31 SFPTORMAT 74 121 SGP FORMAT 5 74 va Ue Pak we o Po va a 36 SHOW CHART TABLE s ooa oues oa eo aa loa ds aen un Fo eve bu Fe 35 SHOW COLOR eee ee ee ee ee ee eene eonun 34 86 99 103 108 SHOW 25 SHOW CONFLICT esi cu vaa uuu ni nn neavnsdesccevsavacende 98 103 107 SHOW 5 94 SHOW CONFLICT WITH SIBLINGS ee ee ee ee ee eene neenon enu 94 SHOW CONTROL SAMPLES IN 30 SHOW DISABLED 5 5 31 SHOW GEL IMAGE ti Fa np ava va doa 29 SHOW INDIVIDUAL 98 103 107 SHOW LADDER SAMPLES IN 30 SHOW LAST EVENT 34 SHOW NAVIGATOR ccecscscscccccccsccccccscscscscccscsccccscscscscscecs 29 SHOW ONLY UNCERTAIN ALLELES eee ee ee ee ee eene nennen enn 67 SHOW REJECTED LOW SCORE ALLELES eee eee eee ee een n 67 SHOW REPORT 5ssssaseesavsssu suse xbv ev ax vaga SED QN RV EE AiR SeS 29 SHOW TOGGLES 34 SHOW TYPE SYMBOL vu va vaa an eu vata va e ovp Eiai 68 SHOW HIDE ICONS icis d eoa Pun bo Ia o au do ae aeo Pro eu
85. C Dye Peak Size Aea Peak Comparison Threshold Icons and Functions F Replicate ComparisonSettings Opens the replicate comparison settings window where the user may determine which qualities of each replicate are compared to one another Peak Compare Items select which quantities to include in the comparison Marker Name and Allele Name are initially selected Peak Match By For convenience some peak comparison items are grouped into three categories Select one of the three categories to automatically select the relevant comparison parameters Peak Comparison Threshold As described below any conflicts arising from the comparison of two or more replicates are flagged Here the user may restrict flagging to only cases in which the difference surpasses a pre set threshold As an example suppose the height of a peak was 1000 RFU and the height of the corresponding peak in a replicate was 1050 RFU If the Max Height Difference was set to 1076 this conflict would not be flagged as it is a difference of only 5 NOTE Peak comparison thresholds will be grayed out unless the corresponding comparison item is selected in the Peak Comparison Items section Edit File Groups Opens the File Name Group Editor allowing the user to group or re group samples within the Replicate Comparison Tool 122 August 2013 Chapter 10 User Management e lx Viewing Options is Use these icons to scroll through dye colors zoom in zoo
86. D THIS CONTRACT CAREFULLY BY USING ALL OR ANY PORTION OF THE SOFTWARE YOU ACCEPT ALL THE TERMS AND CONDITIONS OF THIS AGREEMENT INCLUDING IN PARTICULAR THE LIMITATIONS ON USE CONTAINED IN SECTION 2 TRANSFERABILITY IN SECTION 4 WARRANTY IN SECTION 6 AND 7 LIABILITY IN SECTION 8 YOU AGREE THAT THIS AGREEMENT IS ENFORCEABLE LIKE ANY WRITTEN NEGOTIATED AGREEMENT SIGNED BY YOU IF YOU DO NOT AGREE DO NOT USE THIS SOFTWARE IF YOU ACQUIRED THE SOFTWARE ON TANGIBLE MEDIA e g CD WITHOUT AN OPPORTUNITY TO REVIEW THIS LICENSE AND YOU DO NOT ACCEPT THIS AGREEMENT YOU MAY OBTAIN A REFUND OF THE AMOUNT YOU ORIGINALLY PAID IF YOU A DO NOT USE THE SOFTWARE AND B RETURN IT WITH PROOF OF PAYMENT TO THE LOCATION FROM WHICH IT WAS OBTAINED WITHIN THIRTY 30 DAYS OF THE PURCHASE DATE 1 Definitions Software means a all of the contents ofthe files disk s CD ROM s or other media with which this Agreement is provided including but not limited to i SoftGenetics LLC or third party computer information or software ii digital images stock photographs clip art sounds or other artistic works Stock Files iii related explanatory written materials or files Documentation and iv fonts and b upgrades modified versions updates additions and copies of the Software if any licensed to you by SoftGenetics LLC collectively Updates Use or Using means to access install download copy or otherwise benefit from using the functiona
87. E Fase PvA Y Fuse Tasas ax suo va pu daas 52 MARKER PARAMETERS 53 IVIARKERS i22 inv3aa id an NAE Ia AV EE SERRA FESSVARAVURE FRANE ERAT EX PERSE es 63 MATCH BY FIXED POSITION 4 eee ee eee ee eene eee eene 119 MATCH BY SECTIONS esa en a p basata pn apagar 119 MATCH LADDER sessi 59 135 MATCH SCORE 44 suave visa ve V agen VAM EE UR ad 41 43 44 46 47 MATCHED GROUPS aeu Vua Lan v en ax En 119 xA eura desse faena 93 MAXI OF OPEN vsus opas Va ve ya Ro RERO 29 MAX Se AVERAGE ee vas ea no a vn 60 Max ALLELE LABEL LAYERS eee ee ee eee eene eere rennen enne 30 30 MAXIMUM 53 11 IVIENDELIAN INHERITANCE eee eee ee eene eene sese eese esos 6 6 55 99 IVIENDELIAN INHERITANCE 2cececscscccccccsccccscscscscccscscscsecscsces 93 MENU reki csins 28 MERGE BINS i FEE E 69 MINIMUM COMPUTER 5 MINIMUM HETEROZYGOTE 5 53 MINIMUM HOMOZYGOTE
88. ENTIAL INDIRECT INCIDENTAL DAMAGES OR ANY LOST PROFITS OR LOST SAVINGS EVEN IF A SOFTGENETICS LLC REPRESENTATIVE HAS BEEN ADVISED OF THE POSSIBILITY OF SUCH LOSS DAMAGES CLAIMS OR COSTS OR FOR ANY CLAIM BY ANY THIRD PARTY THE FOREGOING LIMITATIONS AND EXCLUSIONS APPLY TO THE EXTENT PERMITTED BY APPLICABLE LAW IN YOUR JURISDICTION SOFTGENETICS LLC S AGGREGATE LIABILITY AND THAT OF ITS SUPPLIERS UNDER OR IN CONNECTION WITH THIS AGREEMENT SHALL BE LIMITED TO THE AMOUNT PAID FOR THE SOFTWARE IF ANY SoftGenetics LLC is acting on behalf of its suppliers for the purpose of disclaiming excluding and or limiting obligations warranties and liability as provided in this Agreement but in no other respects and for no other purpose For further information please see the jurisdiction specific information at the end of this Agreement if any or contact SoftGenetics LLC 9 Export Rules You agree that the Software will not be shipped transferred or exported into any country or used in any manner prohibited by the United States Export Administration Act or any other export laws restrictions or regulations collectively the Export Laws In addition if the Software is identified as export controlled items under the Export Laws you represent and warrant that you are not a citizen or otherwise located within an embargoed nation including without limitation Iran Iraq Syria Sudan Libya Cuba North Korea and Serbia and that you are not otherwise proh
89. Genetics LLC to provide the information necessary to achieve such operability and SoftGenetics LLC has not made such information available SoftGenetics LLC has the right to impose reasonable conditions and to request a reasonable fee before providing such information Any information supplied by SoftGenetics LLC or obtained by you as permitted hereunder may only be used by you for the purpose described herein and may not be disclosed to any third party or used to create any software which is substantially similar to the expression of the Software Requests for information should be directed to SoftGenetics LLC Trademarks shall be used in accordance with accepted trademark practice including identification of trademarks owners names Trademarks can only be used to identify printed output produced by the Software and such use of any trademark does not give you any rights of ownership in that trademark Except as expressly stated above this Agreement does not grant you any intellectual property rights in the Software 4 Transfer You may not rent lease sublicense or authorize all or any portion of the Software to be copied onto another users computer except as may be expressly permitted herein You may however transfer all your rights to Use the Software to another person or legal entity provided that a you also transfer this Agreement the Software and all other software or hardware bundled or pre installed with the Software including all copi
90. I 2005 C 2431 file Click Save Save Project After a dataset is analyzed and edited the project can be saved as a SoftGenetics GeneMarker HID Project SGF Project files contain the raw unprocessed data files the sample files after processing the process parameters and all edits The project file does not contain any custom or modified Panels or Size Standards To export a custom Panel see Chapter 5 Panel Editor To export a custom Size Standard see Chapter 4 Fragment Sizing Standards NOTE Previous versions of GeneMarker HID saved projects in SFP format This format can be opened with current GeneMarker HID versions dLGeeMeke HD PAri7s eo View Project Applications Tools Help Gb Open Data d a H 9 P E Ud m l Make Nore 9 To save a project go to File Save Project in the Main Analysis window The Save Project box will open Select a directory and enter a project name Click Gel Image Reopen Project fly out menu is selected Click a project from the fly out menu and it will be uploaded to GeneMarker HID Close All E 23 4 33 Save Wr a 6 H Anc da A To re open the project go to File Open Project in the d ze g B PAT 3 Misa 100 120 140 160 180 200 220 240 260 280 300 320 340 360 3 z a 15 Main Analysis window The last folder accessed by qose PAT La
91. ID project sgf sfp to a specified directory Raw data files and analyzed data files with edits are saved within a project file Pull down peaks may result from changes in optical alignment or polymer of the genetic analyzer These data are now included in the traces of saved GeneMarker HID Projects SGF files 28 August 2013 Chapter 3 Main Analysis Overview Close AII Closes a project without exiting the program NOTE It is recommended to select Close All before exiting the program Exit Closes the GeneMarker HID program View Menu The View menu contains options for how the data is displayed in the Main EJ Show Nawigator Analysis window 15 Show Gel Image Show Navigator Gel Image Report H show Report Toggles the Sample File Tree Synthetic Gel Image or Report Table frames open and closed in the Main Analysis window Preference Preference Activates the three tab Preferences box Start up Settings The Start up Settings tab effective only at start up allows you to select the Run Method and General Settings Run Method Classic Appropriate for experienced users The user will move Serio Seis Cw through the program data input settings and display options without prompting by simply following the program s sequential analysis flow Wizard Activates the Run Wizard which will guide the user through the program s operation This setting is best for the inexperienced user
92. IGHTS YOU MAY HAVE ADDITIONAL RIGHTS WHICH VARY FROM JURISDICTION TO JURISDICTION For further warranty information please see the jurisdiction specific information at the end of this Agreement if any or contact SoftGenetics LLC s Customer Support Department 7 DISCLAIMER THE FOREGOING LIMITED WARRANTY STATES THE SOLE AND EXCLUSIVE REMEDIES FOR SOFTGENETICS LLC S OR ITS SUPPLIER S BREACH OF WARRANTY SOFTGENETICS LLC AND ITS SUPPLIERS DO NOT AND CANNOT WARRANT THE PERFORMANCE MERCHANTABILITY OR RESULTS YOU MAY OBTAIN BY USING THE SOFTWARE EXCEPT FOR THE FOREGOING LIMITED WARRANTY AND FOR ANY WARRANTY CONDITION REPRESENTATION OR TERM TO THE EXTENT TO WHICH THE SAME CANNOT OR MAY NOT BE EXCLUDED OR LIMITED BY LAW APPLICABLE TO YOU IN YOUR JURISDICTION SOFTGENETICS LLC AND ITS SUPPLIERS MAKE NO WARRANTIES CONDITIONS REPRESENTATIONS OR TERMS EXPRESS OR IMPLIED WHETHER BY STATUTE COMMON LAW CUSTOM USAGE OR OTHERWISE AS TO ANY OTHER MATTERS INCLUDING BUT NOT LIMITED TO NON INFRINGEMENT OF THIRD PARTY RIGHTS INTEGRATION SATISFACTORY QUALITY OR FITNESS FOR ANY PARTICULAR PURPOSE The provisions of this Section 7 shall survive the termination of this Agreement howsoever caused but this shall not imply or create any continued right to Use the Software after termination of this Agreement 8 LIMITATION OF LIABILITY IN NO EVENT WILL SOFTGENETICS LLC OR ITS SUPPLIERS BE LIABLE TO YOU FOR ANY DAMAGES CLAIMS OR COSTS WHATSOEVER OR ANY CONSEQU
93. ILS appears in the Sample ILS frame Right click in the Sample ILS frame and chose Add Delete or Fix Size to correct size call Right click again and select Update Calibration The changes will be implemented for the sample and the Match Score will be updated When editing is finished close Size Calibration Charts The Size Match Score indicators in the Sample File Tree of the Main Analysis window will be updated Icons and Functions Toolbar Icons Size Calibration Found in the main toolbar of the Main Analysis window View Mode Change the layout of the Calibration Plots frame Adjust the maximum number of rows and columns displayed Maximum number of rows and columns is 5 Chart Synchronize When selected both the Expected Size Standard and Sample ILS traces become synchronized This option is not selected by default Preprocess Raw Data Select Preprocess Raw Data to smooth the samples raw data ILS Auto Fit Y Provides the option to automatically fit the Sample ILS s y axis by the maximum peak height in the trace OR by only the highest matched peaks Print Provides Print Preview and Print of size calibration page providing a physical record of size calibration for the project Save Allows the user to save the size calibration pages as png images Manual Calibration Provides the option of manually entering standard peak sizes if many peaks have been modified This window contains three columns Standard Size fragment s
94. Mixture Analysis 14 0 2015 1 LR 2 x 0 1454 x 0 2015 17 06 Scenario 3 Null Hypothesis Mixture is from A and B Mixture is from A and an unknown person unrelated to B Ihe project contains a mixture and two single source potential contributors Both single source samples are selected and person B is contested In the column for minor contributor allele combinations we see there are three viable allele combinations for the minor contributor person B 10 12 10 14 10 10 or a c b c cc VENENUM INNEN LR 0 1020 x 0 1020 2 x 0 2015 x 0 1020 2 x 0 1454 x 0 1020 1 0 0812 251292 Scenario 4 H Null Hypothesis Mixture is from A and B mu me Mixture is from two unknown people unrelated to A Him oso 2 m os RE OG imo mo x Se and B en aa E t The project contains a mixture and two single source potential contributors Both single source samples are selected and both are contested As in the above scenarios there is only one viable allele combination for person A and there are three viable allele combinations for person B rs ae Likelihood ratio 2 xa x b x cxc 2xbxc 2xaxc LR 1 Qx 0 1454 x 0 2015 x 0 1020 x 0 1020 2 x 0 2015 x 0 1020 2 x 0 1454 x 0 1020 LR 210 245 90 August 2013 Chapter 7 Mixture Analysis 91 August 2013 Chapter 8 Relationship Testing Chapter 8 Relationship Testing Chapter 8 Relationship Testing Pedigree Analysis
95. ON cccccecececscscscscscscscsecs 7 GENERAL SETTINGS os aai a Mad UR Ha Ex aaa aa A UY UR au A dau 29 GENERATION iiu DK USE RE UAE KEEN REAA SURE UE 93 GROUP IDENTIFICATION cccccccececscscscccscscsccccscscscscecs 118 119 GROUPED BY DYE 72 H HETEROZYGOUS IMBALANCE HIM 88 HI HIGH ee eee eee ee nene eee eene eene nennen 26 HIDE EXTRA 5 5 67 HIDE 34 HORIZONTAL MOVEMENT 35 eoa paie eae Rvo ao dr 23 HORIZONTAL REPORT TABLE ececscscccscsccecscscscscscscscsccecscscss 67 I IHE INCONCLUSIVE HETEROZYGOUS eee eee eee ee eene neon 26 26 ILS STATISTICS 45 IMB HETEROZYGOTE 26 55 5 72 IMPORT A PANEL eiissokuekusuts akute aua Ku FoU ORE 16 IMPORT ABI PANELS sccececscscscccscsccccscscscscscscsccecscsces 56 59 IMPORT ABI SIZE STANDARD ccccccccccsccecscscscscccscscsccccscsces 42 August 2013 Index IMPORT DATA FILES sd vea us uk vx Vi VA VR NE WENN V VENERE Na Ya NOVO uM NR EOS 11 IMPORT PANELS 57 usucu ak sua vu Max Maa v eu ea Ra
96. Report OR click the Print Report icon in the Main Analysis window The Print Report options box will appear Select desired settings and click Preview to view the Print Report before printing or click OK to begin printing without previewing the report The reference ladder best match used for each sample and the run date and time for the sample are listed in the first line for each given sample NOTE The View Preference Display Settings options will affect how the Print Report is displayed GeneMarker HID Print Report SoftGenetics Allele Report 7 20 2011 1 40 04 PM GeneMarker HID V2 1 2 Page 1 Allele Report Metro OCME Software GeneNtrker HD v2 1 2 Template Hentifibr single source Project pati 7 no edits SGF Parel Mentifiler Eer tech Tones Sample 1 Ref Ladder PAT Ladder lfsa Fom date amd tire 02 27 2007 23 53 54 gt 02 28 2007 00 29 38 Dye Blue Speaks PAT 1 Cfsa 60 0 100 120 140 160 190 200 220 240 260 290 300 320 340 550 13 DS 12 12 14 29 13 13 Dye Green 8 peaks 1 C fsa 831358 Di35317 0165539 P5133 60 80 100 D 40 160 S m0 D 40 BO 250 oo D 40 x1 10 000 5 000 D 5 E el Dye Yellow 7 peaks PAT_1_Cfsa 60 0 100 120 140 160 180 200 220 240 260 280 300 320 340 360 Dye Red G peaks 1 Cfsa aj 50 1290 60 0 100 120 140 71 August 2013 Chapter 6 Reports and Printing Report Content Options The basic printing options allow the us
97. Setup Phogat Cancel to qut Setup and close ary program you hove ww D Ned lo contre wih the Sobap pagan WARNING Tha progam protected by copyright Uer ard Unaufhonsed producion or the progam cr ans portion ol may ved n cod and omm penes avi wil be proceeded t the meorum aderi possible under lew CE emm xj Er Select Progr am install GeneMarker iecommended imis License Server Manager and License Server Manager Important The korne Serves Manager i needed for oer runrsng bes produci n a von The Licance Server Manager muu ba rutalled on the Server ra na to be voted on Client Computern lt Back Tonine 0s Ho Dry kiy hers bhoan diirai H a bocal licenga was purchased click Register how nerag licor wan prurcheesd dick oniga otak Client Ma previdis n hwark confiqur ean has haan d fect d Hun V alii amp alicn Tima taniere OO x Nin damy kiy has bedr cabira H m bocal licenga was purchased click Registar Hew He licor wen punchami dick Gongs Cieni Cardigans Mrbercuk Client Ma previous n twark confiquriman has ian detected Aegilia Higar Local hey Ery Plage lec Trend Regie Hetero Tend haed bey E Bev caa Chapter 1 Installing GeneMarker HID D C
98. Software provided your backup copy is not installed or used on any computer You may not transfer the rights to a backup copy unless you transfer all rights in the Software as provided under Section 4 2 3 Home Use You as the primary user ofthe computer on which the Software is installed may also install the Software on one of your home computers However the Software may not be used on your home computer at the same time the Software on the primary computer is being used 2 4 Stock Files Unless stated otherwise in the Read Me files associated with the Stock Files which may include specific rights and restrictions with respect to such materials you may display modify reproduce and distribute any of the Stock Files included with the Software However you may not distribute the Stock Files on a stand alone basis i e in circumstances in which the Stock Files constitute the primary value of the product being distributed Stock Files may not be used in the production of libelous defamatory fraudulent lewd obscene or pornographic material or any material that infringes upon any third party intellectual property rights or in any otherwise illegal manner You may not claim any trademark rights in the Stock Files or derivative works thereof 2 5 User acknowledges and agrees that the Software is licensed by SoftGenetics for research use only Any violation of this restriction on use shall constitute a breach of this Agreement User assumes all risk
99. Start End Comments Quality Reasons Bye Size Height Current Value Blue 1438 567 141 3727 045 Desti7313 joo Old Value Blue 1438 567 141 3727 045 D8S1178313 joo Size Height Ht Ratig Area Ar RatiqMarker Allele Differer Quality Score Start Current Value Blue 143 8 567 0 41 3727 045 D85117913 56 7 143 3 144 3 foldValue Bue 438 s jon s po The Print Saved Edit History Report contains the project header with institution User run date time for the project and parameters The table provides a record of each edit time organization user and operation 132 August 2013 Index Index 2 2D OFFSET vastixixdend ia E EIFE XE ICI 36 2 DERIVATIVE TRACE sesssesccsseccssseccssscccsssecssscccssscccsesees 12 A ABI IDEN 56 ABIDE BY PANEL a va Poe Fe ava 72 ACCESS RIGHTS aspis anda Sada eau 129 ACCOUNT AND PASSWORD ae ER EE XE PERF ERR EXEFEVAUE 6 7 8 9 ADD CHILD E M 97 5 94 114 ADD FOLDER Om 11 ADD 94 113 ADD MATE Guccini kd rM C Maa a dd 97 PADD SAMPLES m m 22 ADD SAMPLES TO PROJECT scscscecscsscscscscscscscscscscscsscscscsens 32 ADD USER 129 ADJUSTING veist 57 ADMINISTRATOR 1
100. TE If the user selects Save Size Standard and then answers NO to the Size Standard has been changed save changes the changes will remain in the Expected Size Standard and Size Table until the user chooses Reload or GeneMarker HID program is closed Sample List The Sample List contains a list of all the samples in the dataset Double click the filename and the sample s ILS trace will appear in the Sample ILS frame Use the Up Down Arrow keys to scroll through samples in the list Expected Size Standard and Size Table The Expected Size Standard trame displays as a trace all the known fragment peaks that are expected to appear in the Sample ILS Single left click a green triangle atop a peak to select the peak The green triangle will turn yellow when the peak is selected Additional Options Once a peak is selected right click anywhere in the Expected Size Standard frame The right click menu will appear with the following options Edit Size E The Edit Size box appears Adjust parameters and click OK Size Enter the expected basepair size of the ILS fragment D5Rg Comments Enter free form text regarding the Size I Enabled When selected a 1 will appear in the Expected Size Table Deselect this option to disable the Size in the Size Standard The size standard files GS500 1 has disabled the 250 bp fragment as recommended by the manufacture due to differences in migration of this fragment Disabled size
101. TXT TN ie PY LL and navigate to the saved file s wd AE Ss leiden CTI LE 13 Amm 10 na BAT b CU TIT F 710 13 L M N o P Use the format of the spread sheet i sem s cnim osm cute cns mss stan mei na o ism amd shown here for importing genotypesas txt tab delimited files The AID column has the file name Each marker heading is in duplicate and the extended homozygous format is used SoftGenetics GeneMarker HID for the allele calls If there is a marker 1 of 1 genotypes have been submited successfully with allele drop out or null allele double asterisk should be used in that cell Please see Report Table in Chapter 3 Genotypes saved to the database from a txt file must have the format at follows AID column sample identification Row 1 B The marker names case sensitive repeated as below AA3 f 18 C D E F G H I J K L M N D8S1179 D8S1179 D21S11 D21S11 D7S820 D7S820 5 CSF1PO D3S1358 D3S1358 THO1 135317 D13S317 D16S539 D16S539 D2 Sample_157 11 13 28 32 2 8 10 7 10 15 15 9 3 12 14 10 11 Sample_156 12 13 27 28 7 8 11 15 14 7 12 12 10 11 Sample 155 11 32 2 32 2 8 8 10 10 15 9 3 14 14 10 11 Fe nr 1A 109 August 2013 Chapter 8 Relationship Testing Save and Print Report Find Family Report right click on the report to copy paste directly into an existing docu
102. The Bin Table Report Style offers additional options when a cell in the table is right clicked Insert a Peak at this Bin Site To indicate the presence of a peak at a position when it has been labeled with a Negative Type Symbol right click the cell and select Insert a Peak at this Bin Site The Negative Type Symbol will change to a Positive or Suspected Type Symbol depending on the Quality rank of the peak Hot Key INS Delete To indicate the absence of a peak at a position that has been labeled with a Positive or Suspect Type Symbol right click the peak cell and select Delete The Type Symbol will change to Negative Hot Key DEL Confirm To indicate the peak present at the position is truly a peak right click the peak cell and select Confirm Peaks Only peaks centered within a Panel Bin will change from Suspect Type Symbol to Positive Type Symbol when confirmed Once a peak is confirmed it cannot be unconfirmed only deleted Hot Key CTRL M Delete Bin Columns To completely eliminate an entire column in the Report Table left click any cell within the column then right click the cell and select Delete Bin Columns When Vertical Orientation is selected the Report Table rows which contain the Bin information will be deleted not the columns which contain the sample information Binning Options To adjust which Bins are displayed and to merge Bins in the Report Table 97777 E m click the Bin icon in the toolbar
103. Windows clipboard for pasting into other applications such as Microsoft PowerPoint Save Image Allows the user to save the Synthetic Gel Image as a BMP image file Show in Window Opens a separate window containing the Synthetic Gel Image The separate window can be maximized for closer gel image inspection 23 August 2013 Chapter 3 Main Analysis Overview Image Display Intensity Move the Intensity slide bar located in the upper left corner of a x the Synthetic Gel Image up and down to adjust the intensity ofthe fragments displayed P I Grey Scale a 1 v 1 Go to View Preference Display Settings Gel Image Select T P Gray for Single Dye to change the single dye Synthetic Gel Image l to black and white when only a single dye color is selected when s multiple dye colors are selected the fragments will appear in their SET respective colors Click the Background in White option to a Qn jM Fa ln reverse the black and white exposure for single dye color gel images Electropherogram and Peak Table Features The Electropherogram displays fluorescent signal intensities from capillary electrophoresis instruments as a single line trace for each dye color The signal intensities are recorded in Relative Fluorescent Units RFUs which are plotted along the y axis Along the x axis are the basepair sizes of the fragments The frame
104. Y 099 0005 8113243 Cpu 4 185 0 0062 20 2324 T Copy 094 n 0012 00 mi 074 0 0073 umet Expo T 035 0020 035 MES jos 09 095 10s ng 41 ns amp veim Mine ule 1 pe Mine Lonlipulcr im tatarera Ho d onip selected comburi Hone Average Majai Mx joa z TAT Curnudeeve LA SSE 1 E The file name tree in mixture analysis groups files by the number of contributors When a file under Two Contributors is selected any single source samples from the project that are potential contributors to the mixture are displayed underneath the mixture sample file name Contributor file names are automatically listed in the dropdown menu at the bottom of the screen and peak ratio results are used to determine if the profile is a major or minor contributor Mixture calculation results are displayed in the results table All possible allele combinations are displayed along with the major Mx ratio residual score and major and minor heterozygous imbalance ratios Please see Mixture Analysis Equations section at the end of this chapter for mixture analysis equations Allele combinations that meet all of the mixture analysis parameters are in black font other combinations are in red font 80 August 2013 Chapter 7 Mixture Analysis The Report table on the right side of the mixture analysis screen lists 1 Marker Name
105. a B PE Toc 11 M0Ucase _evadence fia 013531 basi 220 250 Project geram a a E ma S B Summary New 15 samples PC eror 0 1 NC emon 171 Ladder error 0 1 FFatedz 7Hegged 7 Sample File Tree The Sample File Tree of the main analysis window contains two folders The iee x 0000000000 Fie View Project Applications Took Help first is the Raw Data folder which when expanded displays a list of all the 3 dataset samples When a sample is double clicked its preprocessed E Ras Data 3 6 Allele Cal c electropherogram trace will appear in the Raw Data Analysis window See TT Chapter 2 General Procedure B V as aai LM Select Max De seiect All The second folder Allele Call also contains a list of all the samples but Search File when the filename is double clicked the sample s electropherogram trace Sve appears in the Main Analysis window with all sizing information and allele 5 77 ema call filtering applied The Allele Call folder also flags each sample with a Disable ee Edit Comments Mbture green sheet yellow sheet or red strike through indicating size calling success See Chapter 4 Fragment Sizing Standards A red question mark Auto Identify 21 August 2013 Chapter 3 Main Analysis Overview indicates one or more of the analysis parameters were not met for that sample rules were fired Detailed flagging is found in the allele
106. able This indicates a change has been made to that allele Right click any changed allele and select View History The Show Edit History window appears 8 Select a change from the Edit History List to view changes in the Current Old Values table Changes will be highlighted in red 9 To recover a change right click the row in the Edit History List and select Recover Old Value A star will appear in the Recover column 10 Click OK and click Yes when the warning prompts you to confirm R5 S 005 OUI tea 2I Edits History Window ipi xi Current Old Values oye Jee es Ar ato Waker DWerence nus Score Star End Comments usi Reasons Current Value ESTE 143 8 567 3727 0 45 D8S1179 13 143 3 144 3 Old Value 1438 567 3727 0 45 D851179 13 BE 1433 1443 E dit History List EdtTime Organization User Operation Module Recover 2 07 20 2011 13 19 28 Metro OCME Admin Comment Allele AlleleChart 07 20 2011 13 18 03 Metro OCME Confirm Allele AlleleChart amp Print X Cancel Use the Print Preview button in the lower right to Print the Edit History Window or save it as a png or jpeg file 131 August 2013 Index SoftGenetics Edit History 7 20 2011 1 20 44 PM GeneMarker HID v2 1 2 Page 1 Run date and time 02 27 2007 23 17 55 gt 02 27 2007 23 53 36 Ht Ratid Area Ar_RatiqMarker Allele DiffererQuality Score
107. ack up the database files on a regular basis Procedure 1 Open data file or previously saved project 2 Run Wizard to call alleles 3 Select Relationship Testing from the Applications drop down menu 4 Select Tools Allele Frequency of the appropriate population from the Tools drop down menu 5 Select Tools Genetic Analysis Settings if settings other than the defaults are desired Individual sample 1 Tools gt family group tool gt OK to allow selection of each individual in the file as a separate node 2 Use the Family dropdown menu to select the individual file 3 Right click on the node and select find family from the drop down menu 4 Left click on the Report icon in the tool bar to display the file name and any duplicates of that file found in the data base The samples with the highest LR for each relationship type are displayed in descending order in addition to the sex number of matched alleles and matched markers Relationship Testing Sertings gt amp Family 1 23 Samples All Markers Search d f Ad Eae Samples sie 2 Samples A Charts Report B Calculation Details hirik ees Feeyaeric y Fy blaar TA 7 Same Individual TT Select Node PAT_1_C fsa 1 24E 17 Paii 05 Harari Deselect Node Father Son be Mutation PAT 1 F fsa 2 94E 04 Muir Rate am Select Parents Mother D aughter NUPTIAE JOE 2E 1 7 17E 03 Select Sibling PAT_1_M fsa 5 ff Step Diference Select Fa
108. ailable auto fit the Y axis using peak intensities of the alleles or the user can select the ranges for the X and Y axis Browse by AII Colors Displays a comparative view of sample electropherograms by dye color Individual samples can be selected from the drop down menu m m 19 Importing Population Specific Allele Frequency and Mutation Rate Information The Relationship Testing application in GeneMarker HID has the allele frequencies for major USA populations and the mutation rates specified by AABB pre loaded Customization with population specific allele frequency and mutation rates is easily accomplished using the open folder icon of the allele frequency and mutation rate tools Population specific allele frequency and mutation rate tables must follow the format of preloaded files and be saved as a txt tab delimited file Tools gt Allele Frequency pg gt gt Save 108 August 2013 Chapter 8 Relationship Testing Building the Database DataBase gt Save to DataBase and select from three different routes 1 Directly from the current project pcs Se iu cmm BAT t Fh Adelie DO Y 3 5 3 gl 3 Larikan TE Submit ww tw PAT Hderi he CUE 73 Beas MmU T DEE COVE 13 ma m TI Em 2 From a CMF file select Load from cus mde um CMF and navigate to the saved file s zt ema tira 1 13 Xr lir 15 11 me V TE i x 13 wiz 3 Froma TXT file select Load from
109. alysis Overview Chapter 3 Main Analysis Overview Chapter 3 Main Analysis Overview Main Analysis Window Menu Options Main Toolbar Icons Additional Analysis Options 20 August 2013 Chapter 3 Main Analysis Overview Main Analysis Window The main window of GeneMarker HID has an easy to use layout The sample files are displayed on the left the Synthetic Gel Image is displayed at the top Electropherograms appear below the gel image and the Report Table is on the right side of the window To resize the frames in the Main Analysis window simply place the cursor over the partitions that separate the Synthetic Gel Image Electropherogram Sample File Tree Report Table The cursor will change to a two headed arrow bisected by two vertical lines Hold down the left mouse button and drag the gray vertical line in the direction you wish To open and close the frames use the Show Hide icons in the main toolbar Main Analysis Window Sample File Tree Report Table oynthetic Gel Image Electropherogram Gesch ve eei File Yew PASCO Apphcatinons Jools Hee NERT BH 4A4VW Z5 b i uefi 5 zs I Is H Report 8 Alek Cal LASSI 4 v a Mi Dhcasel _eviderce_longenwyer a i enone tuned a quocp i s mtm he D MbWScare _evidence Jg BB ooa mirer longs d victimis x qaaa D Hoad victimis LO PorciPlexTE LADDER fra g 5 B NC Posen Tine ita
110. as potential mixture MX 78 August 2013 Chapter 7 Mixture Analysis Mature Anatyics Seting Mixture Analysis we SU e vere Mew em Macher Markers 12 E Pes Heg Pea Area 1 Select Applications gt Mixture Analysis D BE See k Rew Rao x Ainin Fequerc Populaisin 2 Launch Mixture Analysis Settings Ce ge utem 1 040 E 0 50 t 2000 060 gt pude LR Value of 0 in Cuma sitve LA Analysis Type Peak Height or Area e bees Tr Minimum HIM heterozygous imbalance rm ei Sin end minimum is on a sliding scale dependent on the lio Dopad lt 100 Range Right click to edit Min HIM parameters Maximum Residual we infer that the se El likelihood of the peak areas given the combination of genotypes is high if the residual is low P Gill et al 1998 is calculated as gt va observed pa expected using peak areas and mean Mx to compare the observed proportions to the expected proportions Minimum Major Weight parameter allows selection of potential combinations based on the Mixture Proportion Mx Set minimum stutter ratio and drop out intensity In cases where a valid allele combination would result with allele drop out a Q is used to represent a possible un amplified allele The value set in the parameter for allele drop out is used in the HIM and Mx calculations For example if the genotype of a mixture
111. ate produced any allele calls at a given marker the status column is filled with an N for Null Final Genotypes Column The final genotypes column displays the consensus genotype for each marker of each replicate group 123 August 2013 Chapter 10 User Management Concordant Case If the status column is filled with a C the final genotype column is Report 3 automatically filled with the genotype of both E replicates pae e Null Case If the status column is filled with o pe an N the final genotype column is THO automatically filled with the genotype of the 251 replicate which has allele calls D18551 Allele1 Allele2 Allele1 Allele2 w wor eA Penta E D55818 013531 D75820 D165539 CSFIPO In this case it is up to the user to manually Pata determine the final genotype This can be o pe done using a dropdown menu in the ooo WA corresponding cell of the Final Genotypes column es Discordant Case If the status column is filled with a D the Final Genotypes column is left blank FGA t N t p t t t t t t t t t t t t t t From the dropdown menu select the o pP genotype for the entire locus The dropdown o menu can be accessed from any of the final le ue genotype cells in the same row as the Marker Allele2 Allele Allele2 Alleei Allele discordant marker After selecting a genotype 17 17 the status will change to E
112. be included in the replicate comparison window 126 August 2013 dl GeneMarker HID Untitled File View Project Applications Tools Help 5 gt m mms Untitled H Raw Data Elf Allele Call EE B 1 B LD 57 Alleli 27 B 50 020410 140 3 B 5idtem 1 a C 3 i 1 b D 3 iB 544tem 1 abc 3 LA 5tem 1c E 4 B 554tem 5 a G 4 i B 5b tem 5 b H 4 B B Htem 5 abc 4 SHtem Sc C 5 58 020410 140 B H Bl tem B a E B Bp 624tem_6_b Fl BE B E3ltem B c G B B amp 4 tem abc Select Page Select Mext Group Select Max De select All Search File Ctri F Sample Info set Sample Type Sort Samples Disable Edit Comments Chapter 10 User Management 127 August 2013 Chapter 10 User Management Chapter 10 User Management Chapter 10 User Management Procedure User Manager History Settings Edit History Audit Trail 128 August 2013 Chapter 10 User Management Overview User management may be implemented after installation of GeneMarker HID The administrator activates User Management from the Help drop down menu User management provides control of user access rights and automatically generates an audit trail of all edits EL Setup Administrator User Manage History Sera Proc e dure Uss Nas Use Type Tire EV 1 Select Help User Management T 2 The Login box appears 3 Click Run User Protection to activate the setup D ol Admini
113. cecececscscscscccscsccecscscscccscees 42 SAVE CHANGES osoxescex svn Sek 43 59 SAVE CHANGES WITH 5 59 SAVE PEAK TABLE avete uet daba a atu daa da Ua Maus aqua UR a 26 35 SAVE PEDIGREE FILE eee ee ee een 97 98 103 107 SAVE PEDIGREE TREE 5 55 soupape 97 SAVE PROJECT 28 74 SAVE REPORT avat eva tu vat evel 35 SAVE REPORT TABLE viveievacevesccaveveveduseceveteteeuseOaseveseduseceses 28 SAVE RUN 5 31 SCF osos tva an Ha a Rut 121 SCORE asana aooaaeoa aE 25 SECOND DERIVATIVE TRACE ee eee eee eee eee eene nene tope enne 15 SEGREGATION ina sana AVR OR VEA VERAM EY VE a E Fas 93 SEGREGATION eaii iNi 99 SELECT MARKERS iva us anag vaesieonus vsvsieusled pss vaveessescsavawsusis 98 SELECT NODE sooo vasa ve vena nha vat a an pauta ca ain su en pa Uta ena aqaa ro eun ph 93 SELECT DESELECT INDIVIDUAL 93 SERVER COMPUTER vss wseeysd enekezesvsesosoosdep deos qupE Use oaae 7 SET AXIS Cu 35 99 103 108 SET CHANNELS EYE ER 31 SET SAMPLE TYPE 2
114. cs License Server 11 Click Next in the Start Installation window to install License Server Manager 12 Select the Launch License Server Manager option and click Finish Unam pore rimo uon tn r Pes program x ary piton of i m y result in bira Cred and coral penalbes and vell Be pecpescut sd 19 Psi redpornuns doen pobtb s undis lia Lie e Server Xy Sol erehes Loerie Server hac been successfully naaed Dick the Frah button 1o eset thee rataiahon Launch License Server Manager 13 Click OK in the Install window to restart the system 14 The Installation Wizard will close and the system will restart compte Press Conclto tunto Vind 3 without restarting 15 Eject the SoftGenetics CD Cancel Im M Register License Server Manager for GeneMarker HID Usage E 5 SoftGenetics License Server x 1 Open License Server from the System or Icon Tray by clicking the LSM This program has not been registered ICOn To complete the registration process click on Help and select Register Note A red star indicates the License server is not running The icon with a white star indicates the License Server is running properly 2 Click OK in the dialog box to proceed with registering License Server from the License Server Manager console 3 Select Register from the Help menu to activate the Register E Product window an Regie Dirdre Onine Reginan 4 Select GeneMarker HID
115. d custom Panels saved to the Panels folder in the SoftGenetics GeneMarker HID directory Single left click on the panel name to display the trace Single right click on the Panel name Delete to displaythe pop up menu of options Double click the Panel name to expand Export the folder and view the Markers associated with the Panel Single left click the Reload Marker name to display that Marker in the Overlay Trace frame Set As Project Panel Additional Options To see additional options for each Panel right click the Panel name and the right click menu will appear with the following options Edit Panel EN Panel Parameters Panel Name Profiler_Plus Ploidy 10 Decaploid X 51 August 2013 Cancel Chapter 5 Panel Editor Edit Panel Opens the Edit Panel box Editing the Panel Name field will change how the Panel is labeled in Panel Editor Set the Ploidy from Monoploid 1 to Decaploid 10 to Unlimited If the number of peaks within a Marker exceeds the Ploidy setting the additional peaks will be labeled Off Ladder OL and given the Undetermined red Quality rank and PL Quality Reasoning When Unlimited is selected the PL rule is never fired Delete Panel Select Delete Hot Key DEL to delete the Panel from the Panel List and from the SoftGenetics GeneMarker HID directory NOTE This action is irreversible Export Panel Opens the Save As window Choose a directory folder and click Save The Pa
116. d in red Clicking on the marker in Tx nu ue the list links to the section of the ws ade hem electropherogram where the conflict can be D s1128 Ium visualized CNET sho o monde nas ase 16 16 028718 THI 5 3 3 L 0138317 yog pisss 12 ii pm 74 4 0188433 142 15 WWA E 14 TPOX 12 6 518551 15 12 AMEL Y x 59818 a 1 FGA 14 M 111 August 2013 Chapter 8 Relationship Testing Renaming Tool If the file names are not adequate for use in the family group editor the renaming tool provides transient naming conventions and allows the user to use the automated pedigree drawing instead of manually diagramming the pedigree Procedure Iz 1 Click on the icon to activate the renaming tool z 2 Enter values that have a family identifier file_relationship field RR 3 C Child F Father M Mother for several children T in a family group number them C1 C2 C3 ei iun 4 Enter Group Identification and Control Identification j values as previously described for the family mens m Grouping Tool vemm 5 Click Match and then OK to draw the pedigrees REN 6 Right mouse click on the parent node select Edit vso ur Node he 1 7 Click the contested box to obtain the PI calculation NE caen results using the trio or motherless case PI equations from AABB Recommendations for Relationship Testing
117. ds To open the Size Template Editor select Tools Size Template Editor from the menu bar OR click the Size Template Editor icon in the Run Wizard Template Selection box Due to differential fragment mobility in capillary gel electrophoresis a sizing standard must be applied Each sample run through a CE instrument will contain an Internal Lane Standard ILS The ILS contains peaks of known size and is usually tagged with red or orange fluorescent dye Since the ILS dye labeled fragments migrate through the same capillary as the other dye labeled sample fragments they are subject to the same environmental conditions and can therefore be used as a guide to determine the size of the other fragments in the sample A Size Standard template is applied to each ILS and sizes between the known ILS peaks are interpolated NOTE GeneMarker HID is optimized to size fragments with linear mobility Larger fragments or those run through a high viscosity gel i e agarose do not migrate linearly and therefore cannot be analyzed with GeneMarker HID at this time Size Template Editor Size Standard List Expected Size Standard sample List Expected Size Table Sample ILS pn EX sse lerne late n P D m E og See Starches a Ba ET400A ETAWA 55 100 2 amp n 65 050 Gs 200 5535 G55 S5 1 bs GS ETKO frg CO HOT tea Math Wdadadddadddad E amples af frag O01 HU isa na
118. e peaks with Quality ranks of Check yellow 0 t 9 2c 202 3 eer cone and Undetermined red praseza 318 6 1021 8787 27 0 1051 9178 Show Rejected Low Score Alleles When selected the a ee IR 00 Jpuesiood 120 1155 605 3714 BH 135 135 1 407 3003 1389 14114 587 1526 peaks with peak scores below the Run Wizard rss 16s sem a 8 Additional Settings Allele Evaluation Peak Score Reject cL o 5555 p18ssae 350 8 es 8 setting will be displayed in the table 446 2 E ie 4 52 E Hide Extra Sample Names When data is displayed in pas 312 5 3963 2 8 H 2 99 ros scr 215149 118 0 39306 Vertical Orientation the sample names are repeated for mg 223 5 51755 each row of data that the sample is associated with If 202 2 E ise 1562 52108 4906 228 7 5804 4341 7100 Hide Extra Sample Names is selected then the sample ssv2 m 2 2544 35036 name will only appear once in the first of the rows itis Lss associated with Allele Count Allele Report Settings The Allele Count Report Style displays the number of alleles present in are the Panel Marker Sample names are listed in rows in the left column M Allele List and Markers are listed along the top row in columns A Total Number Forensics column lists the number of peaks detected in the sample C B
119. e the Trace Data to toggle from the i Ji LR report to the traces Use the Show all tn E 00 combinations toggle to display either all Ei ww ur jm wir Brame genotypes black font for combinations that Stree Tx Uer oe a a conform to all of the mixture analysis parameters and red font the combinations that c oe Bassas sagt EAEE SSB SERYRFRRnaR m empeoceocnmsucnmcimoemmemmmoemnaogssumnceecooogd tEukgHuuvEHZczHSBSERSZ2SYT 2292888222235 385gERETEI do not conform with the mixture analysis LN odi parameters OR toggle to display only the y Li he ee black font Submit a reference profile to the Relationship Testing Database A profile from a reference sample can be submitted directly to the Relationship Testing Database from 1 gs this mixture analysis screen eee 4 amp ng Mick BOT ha Joi MigF HOS isa x n F TE 1 Use the Submit Genotypes to 81 August 2013 Chapter 7 Mixture Analysis Database icon 2 Select the appropriate file s 3 Submit 4 OK Search Relationship Testing Database for exact matches amp ai 1 Use the Search Database icon to search the database TW ER Search Minar for exact matches Search Unknown Contino Select the desired profile from the drop down menu
120. ed Size Standard for the dataset If the Size Standard name is known simply single left click the Size Standard name in the Size Standard List and click OK in Size Template Editor The selected Size Standard will then appear in the Size Standard field of Run Wizard Template Selection box and will be used to size the data Alternatively if the Size Standard name is not known follow the Best Match steps below 1 In Size Template Editor select BestMatch Match All 2 The Data Processing box appears Eye 3 GeneMarker HID cycles through all Size Standards 4 Click OK when Data Process is finished fem i as 5 The Size Standard with the best average Match Score across all samples in the dataset will be highlighted in the Size Standard List and appear in 5557 the Expected Size Standard frame m NOTE BestMatch will not always choose the correct Size Standard User inspection is required pe 6 Once the Size Standard is chosen click OK in the Size Template Editor _ 7 The selected Size Standard will then appear in the Size Standard field of TF Run Wizard Template Selection box and will be used to size the data Custom Size Standard Creation Follow the steps below to create a new Size Standard based on the dataset currently uploaded to GeneMarker HID Input Dialog Tes New Size Standard Name Test Size Standard 41 Cancel August 2013 Chapter 4 Fragment Sizin
121. ed by the dye colored brackets at the top and bottom of the electropherogram NOTE Only in the Panel Editor do the vertical gray bars within the electropherogram indicate the center of the Bin For all other views in GeneMarker HID the vertical gray bars in the electropherogram indicate the center of the detected peak Panel Editor Panel Table Trace Overlay Project Panel Panel Templates BU LM DUET g Wir me CT IT LL PT e T Corea e Sample List 32 o5 oomonmonoouooooc SSRRSHRSS S amp S SHSFRAARSASS ITEEEFEEEEEEETE GRO ij gt lp svestssesseovcsategsweg zT z ZA an 0 LLELLE 2 i rxzroontT327702797972500 5 5 5 sae x Sonno y 7 11 r1 53 2582 52 58 5 111 1614 1 Project Panel The Project Panel list includes the template panel used for the project and all allelic ladder samples that fit the pattern recognition to the selected template panel These ladder file names are in bold font in the main analysis window when Select Best Ladder is used for the Run Wizard GeneMarker HID selects the best fit ladder from this list as the reference ladder for each sample in the project The reference ladder for a given sample is listed on the print report Panel Templates The Panel Template List includes a list of all pre defined an
122. ed samples will be sized and the ee eae allele calls will be filtered according to the parameters set in the Run Wizard 22 August 2013 Chapter 3 Main Analysis Overview Sample Grouping From the main toolbar select Project Apply Sample Grouping The File Name Group Editor tool will appear See Chapter 8 Additional Tools Select Group and Control identifiers and click Match Click OK to apply the matched groups Group numbers will appear next to the filenames in the Sample File Tree Use the Select Next Group right click menu option OR CTRL PageUp Down to open samples in a group To disable the Sample Grouping feature go to View Preference Others and uncheck Enable Sample Grouping Synthetic Gel Image and Electropherogram with Peak Table The Synthetic Gel Image and Electropherogram displays are associated in the Main Analysis window Both display the fragment information in a visual form When GeneMarker HID is initially launched all dye colors are displayed in the Synthetic Gel Image and Electropherogram at once Single left click the Show Color icon in the main toolbar to cycle through the dye colors or use the Show Color drop down menu to disable individual colors or Show Hide All colors Navigation Zoom In Out UTER s In the Synthetic Gel Image or the Electropherogram hold down 575 OSS wsi aal the left mouse button and drag a box from upper left to Get image lower right around the area you
123. eile RUE 7 fannie 3 File name s of matching profiles will be added to the drop down menu 4 LRsare displayed in the Report Table N Before search database icon the drop down list has only the single source reference files listed After Selecting the profile to search the database all files that match are displayed in the drop down menu M y earar Donius z UE FOIT aj Fade Oe iai O Fan FOIT 14 4 immi Ammam Hie Pe Dinan 1 5 Select the file to display the LR results in the Report Table What to Expect Without a Reference File In some cases only the mixture sample and the victim reference sample are available For kss some example DNA isolation from rape cases often 72752 E EU ERES has successful male female cell separation that E n i Hus du umm provides a mixture where the perpetrator is the usc CE 5 HI SS dm dm major contributor and the victim is the minor RES NOSE e 8 D 5 contributor Results shown in the following 3 4 fie ge figure allow the analyst to identify a single Hz ss 55 profile that corresponds to the perpetrator in be the Major Contributor column This profile can T MGR mtem be used to epe 5 1 Search the database in GeneMarker for an exact match or match right from the mixture analysis screen 2 Search for potential relative see Chapter 8 section Data Base Search Finding Nearest Relatives 3 Save and
124. ency Mutation Rate specified by AABB aene Population Statistics for the family groups under analysis Kinship Analysis 2pq heterozygotes and p p 1 p 0 where 0 0 01 homozygotes Mii ies poke Genetic Analysis Settings allows setting the above options at the same time 102 August 2013 Chapter 8 Relationship Testing ni Ss so R e amp New Pedigree File Select New Pedigree File to create a new pedigree with multiple families OR select New Family to add a family to the pedigree file Enter the first family member s information into the New Family New Individual box and click OK to create a new Pedigree Tree Open Pedigree File Launches the Load Pedigree File box Select a PED or PRE file to upload the SMP file will automatically upload and click OK Save Pedigree File Launches the Save Pedigree File box Enter filename and change directory to save the Pedigree Files PRE SMP DAT Show Individual Name When selected the individual ID will be displayed in the nodes of the Pedigree Tree Update Sample Data Select to refresh the Mendelian inheritance calculation after a node or allele is edited and after selection a different family when the show genotype display is used Relationship Testing Parameters Launches the Relationship Testing Settings box Options for selected samples or all samples selecting the appropriate allele frequency for the population mutation rate and prior probability gender known a
125. ent Computer sees rne nnne naa 9 Upgrade of License Server Mondger isses sisse sweets eee rv a aeia a 9 Upgrade of GeneMarker HID Software on Client Computer lisse eene enne nennen annees nnns 9 Gia cm 9 CHAPTER 2 GENERAL PROCEDURE sss a NER RDRERSRE RENE S TER EVERY aUa Da ud T ETANERR SS URN S RSS M RR OK MA EN V eR 10 IMPORT PAY eic 11 11 FO TCS E A 11 RAW DATAA IRL e MO 11 MOn TODO ICON NETTE 12 to EE EERE AEAEE E EEEE EE ETE EESE 13 PROCE S TA 15 T O ES E T OT rE OTE NET 15 KUN Wara DOLAS OCC SS ot E EEE ERE E EEEO 16 Run Wizard Additional SCTUNGS usus saei nrnna ren Prpi etam nca Fat Puertos they t Parte ciru tS A ATOA 18 ADJUST ANALYSIS PARAMETERS catt qus dna Ex 19 Re analyze with Run Wizard eerte 19 Re analyze with Auto saa assess saa ases
126. epair of the Bin For use only with panels that do not contain variant alleles 0 in the control column Ame R 5 Saving a panel with signal information For optimal automatic panel adjustment a panel should be adjusted manually the first time and then saved with signal information from several ladder samples from the same genetic analyzer and used for analysis of any samples from that analyzer To manually adjust alignment hold down the shift key and use the mouse to shift all of the bins for a marker with the entire gray horizontal bar containing the marker name or to shift individual bins hold down the shift and use the mouse to move the vertical bar into place This signal information is used to adjust to any normal variation of different runs from the analyzer p 7 wu 7h eee T 4 C4444V CCCCCRS i e 8 For example if the laboratory routinely uses two 3130 genetic analyzers 3130 1 and 3130 2 there would be a panel saved with ladder signal from each genetic analyzer Saving the panel with signal information from the ladders provides the program with the information needed to automatically adjust the panel with subsequent data A common occurrence is a difference due to run time variation causing a shift in the peaks of 1 5 base pairs The major panel adjustment icon can be used to automatically align the panel to the current ladders 60 WI
127. er to choose the Print Type Samples to print Dyes to include and Content options Each electropherogram will be automatically labeled with its respective sample file name in the printed report The Advanced button provides more options Print Report Print Type Beg C Chart Overlay Normal All Print Report options are available when Normal Print Type is sna ER selected C Al Samples Del ve Chart Overlay Prints only the Electropherogram with the report irl d Samples IV Electropherogram fV Dyed All Samples Prints all the samples in the project Advance Selected Samples Prints only those sample files that have been selected in the oe V Hide Bins Main Analysis window Sample File Tree Contents Bor Electropherogram Prints the peak trace for each dye color and sample selected NOTE The zoom setting of the Electropherogram in the Main Analysis window will be represented in the Print Report Zoom out fully to include all peaks in the Print Report Peak Table Prints the Peak Table for each dye color below the dye color s electropherogram trace NOTE If neither Electropherogram nor Peak Table were selected the Print Report will contain a list of each dye color selected for each sample selected and the allele count within each dye color Forensic Table Prints the allele calls genotype table on the final report Dyes Dye 1 6 Click the checkbox to include the dye co
128. ericans hispanics and other populations of the United States of America and Brazil published erratum appears in J Forensic Sci 2001 Nov 46 6 J Forensic Sci 46 736 61 http www promega com applications hmnid referenceinformation popstat custstat Allelefreg htm CODIS 13 2001 Budowle et al Budowle B Shea B Niezgoda S Chakraborty R CODIS STR loci data from 41 sample populations J Forensic Sci 2001 46 3 453 489 http projects nfstc org workshops resources literature CODIS9620STR9620Loci9620Data9620from9620419620S ample pdf African American US Caucasian and Hispanic from the FBI column in Budowle 2001 were previously published in Budowle B Moretti TR Baumstark AL Defenbaugh DA Keys KM Population data on the thirteen CODIS core short tandem repeat loci in African Americans U S Caucasians Hispanics Bahamians Jamaicans and Trinidadians J Forensic Sci 1999 44 1277 86 106 August 2013 Chapter 8 Relationship Testing ESX ESI 2010 Hill et al Carolyn R Hill David L Duewer Margaret C Kline Cynthia J Sprecher Robert S McLaren Dawn R Rabbach Benjamin E Krenke Martin G Ensenberger Patricia M Fulmer Douglas R Storts John M Butler Concordance and population studies along with stutter and peak height ratio analysis for the PowerPlex ESX 17 and ESI 17 Systems FSI Genetics April 2010 http www cstl nist gov strbase NISTpop htm http www fsigenetics com article S1872 49 73 10 00071 2 abstract
129. es Updates and prior versions and all copies of font software converted into other formats to such person or entity b you retain no copies including backups and copies stored on a computer and c the receiving party accepts the terms and conditions of this Agreement and any other terms and conditions upon which you legally purchased a license to the Software Notwithstanding the foregoing you may not transfer education pre release or not for resale copies of the Software 5 Multiple Environment Software Multiple Language Software Dual Media Software Multiple Copies Bundles Updates If the Software supports multiple platforms or languages if you receive the Software on multiple media if you otherwise receive multiple copies of the Software or if you received the Software bundled with other software the total number of your computers on which all versions of the Software are installed may not exceed the Permitted Number You may not rent lease sublicense lend or transfer any versions or copies of such Software you do not Use If the Software is an Update to a previous version of the Software you must possess a valid license to such previous version in order to Use the Update You may continue to Use the previous version of the Software on your computer after you receive the Update to assist you in the transition to the Update provided that the Update and the previous version are installed on the same computer the previous version
130. es application of a sizing standard filtering of noisy peaks and comparison to a known allelic Panel GeneMarker HID combines all these steps in one simple tool called the Run Wizard To access the Run Wizard simply click the Run Project icon in the main toolbar Run Wizard Template Selection T Template Selection Procedure as l 1 Click the Run Project icon in the toolbar 7 Seed ang iei i emi cr 2 The Run Wizard Template Selection dialog box will appear Kae lt a 3 Select a template a previously saved set of size standard m XD a standard color and analysis type named for future use OR k REM sinici Bue 3 select a new combination of size standard standard color and Andi Tyee i H analysis type T 4 Click Next when finished A um Icons and Functions Template Name Select from existing pre made templates or create your own by entering a Template Name and clicking the Save button When the settings for all sections of the template have been completed Template Selection Data Process and Additional Settings use the Back button to return to the Template selection screen and then select Save Note only individuals with access rights to change analysis parameters may save Run Wizard Templates To create a new template click Select an existing template or create one A template can also be selected from the list of available templates in the left section of the window and then
131. essa asses 19 he oanalvaze Individual Samples csse mex i p a n d DEL M ga NEUE 19 CHAPTER 3 MAIN ANALYSIS OVERVIEW 20 MAIN ANALYSE WINDOW mc 21 SIO FUE TCS eet den oe ease wie umts advan aed ae NNNM eee 21 Synthetic Gel Image and Electropherogram with Peak Table isses eene 23 REPOT cT Oy H N 27 re T M 28 uL mM 28 I UII M WO 29 gives 31 VAN a kom MENU C 32 ore IVC arate T m 33 34 d doct 34 P ODITONALANAO SIE 36 Browse 36 Prolile Compar VIEW alae 36 CHAPTER 4 ERAGIVIENT SIZING STANDARDS
132. et directory before installing the newer software Installing into a New Directory If you choose to install the newer version in a different location be sure to specify a unique directory name or Program Manager Group for the upgrade to prevent overwriting any previous versions of GeneMarker HID Several files created by the users or created during analyses conducted by the previous version of GeneMarker HID will not be recognized by the new version of GeneMarker HID unless they can be found in the directory of installation If you intend for the new version to recognize these files then you will need to copy them from the older version s installation folder and paste them in the folder containing the new version of GeneMarker HID Some of the more common customized GeneMarker HID files are GeneDB mdb GeneMarker HID mdb codis ini CommentsTemplate ini ExpTemplates ini Panel folder and SizeStd folder Upgrade Procedure Text based 1 Before proceeding prepare a backup copy of the previous version of GeneMarker HID recommended Double click the GeneMarker HID executable file EXE on the SoftGenetics Upgrade CD Proceed through the Installation Wizard as described in the Installation section above Once the Installation Wizard is complete launch GeneMarker HID by double clicking the new GeneMarker HID desktop icon OR open the Start menu and navigate to SoftGenetics GeneMarker HID the version that was just installed GeneMarker HID pr
133. et in the Forensic Tab of Preferences Automatically Re Sort Report jw Automatically Scroll Chart to Alleles When Selected in Report Show Disabled Samples in Report Open Mutiple Charts When Browsing Report Others The Others tab has additional selections for the project Enable Sample Grouping When Project Apply Sample Grouping is implemented the Enable Sample Grouping option will be automatically selected De select Enable Sample Grouping to inactivate the Apply Sample Grouping option The Apply Sample Grouping information is saved and can be recalled by selecting Enable Sample Grouping See Chapter 8 Additional Tools Filename Group Editor Gabe nass Pics see o ies Automatically Save Run Wizard Parameters to INI file Att Automatically saves the Run Wizard parameters in an ini file when the project is saved The location is the same location as the saved project and the name of this file is the name given to the SaveProjectName RunWizardParameters ini Channels Opens the Set Channels box and allows the user to choose from ABI MegaBACE and Beckman Coulter standard dye colors The user can also manually enter dye SHA Snes color and name The default channel color setup is ABI See Chapter 2 General Procedure Preferences Automatically Save Run Wizard Parameters to INI file Hi Channels b Run Project Menu J gt Auto Run The Project menu contains options for ho
134. export for use with external database searches 82 August 2013 Chapter 7 Mixture Analysis Submit a profile from deconvolution of mixture sample to the Relationship Testing Database 1 Toggle to display only the allele combinations amp that conform to the mixture analysis parameters ead Submit Major 2 Review to confirm that a single profile has Submit Minor resulted for the major contributor 3 Select Submit Major Submit and OK NOTE Please back up the Database file on a regular basis 51 SeneDB mdb El GeneMmarker_HID mdb Fall GeneDB Idb Gal GeneMarker_HID Idb Search Relationship Testing Database for matching profiles amp i 1 Use the Search Database icon to search the database for exact Jr matches PAG Wien E 2 Selectthe desired profile from the drop down menu T Cis l EHE 3 File name s of matching profiles will be added to the drop down menu 4 LRsare displayed in the Report Table Contributor 1 MixD5case2 vicimMmo IV Before using the search database icon the drop down list has only 1r the single source reference files listed Average Major M 0870 Cumiive a VES After selecting the profile to search the database all files that match are amp 4 displayed in the drop down menu These results indicate that there is an sido Rust e individual XYZ in the database that matches the major contributor in this mixture in addition
135. f the expected phenotype The node is filled in white Affected The individual expresses the phenotype The node is filled in with diagonal hashed lines Contested Select Contested to activate the ability to identify a file as an alleged father unidentified human remains UHR or a contested child Sample File Select the individual s sample file from the drop down list Only samples in the current dataset will be available If no sample file is chosen the node will be grayed out Additionally drag and drop a sample from the Sample List onto a node to associate the sample with the individual node From Database Select the individual s sample file from the drop down list Any sample that was previously saved to the database will be available Add Family Members To add family members to the Pedigree Tree right click a node and select Add Mate or Add Child The Add Mate or Add Child box will appear Enter the individual s information and click OK Save Report Export Bitmap to save the diagram 4 114 August 2013 Chapter 8 Relationship Testing Paternity Index Calculations Overview Paternity index PI is calculated using the Recommendations of AABB Standards For Relationship Testing Laboratories Appendix 8 The combined PI is displayed to the right of the file of the alleged father and the Calculation Details tab contains the results for each marker As with t
136. file names Loci Description File DAT Contains gene frequency information generated in the Pedigree Parameters box There are two ways to begin a relationship test with the Pedigree tool upload a previously created pedigree file OR create an entirely new pedigree file The procedures are described below Mil Load Pedigree File Upload Previously Created Pedigree ET 1 In Pedigree tool click the Open Pedigree File icon j PAUsers SoftGienetics Desktop TestPed pre 25 2 TheLoadP edig ree File box appears v IndividusSample Accordance File SMP V Auto Load 3 Click the Open Files icon next to the Pedigree Files field Vener 4 se di Windows Explorer window to locate the Pedigree File PED on 5 Select the file and click Open NOTE The Pedigree File selected must be associated with the samples currently uploaded to GeneMarker HID If the sample filenames are not the same the Pedigree Tree will appear grey and will not link with the Sample List or Electropherogram Chart 6 The directory path will appear in the Pedigree File field 7 The same directory path and filename will appear in the Individual Sample Accordance File field if the SMP file is located in the same folder 8 If the SMP file is located in a different directory click the Open Files icon next to the Individual Sample Accordance File field select the correct directory and click Open 9 Click OK in the Load Pedigree File box
137. for Edit to reflect E a ee i that the locus has been manually changed mic s ONU MS aa The user also has the option of selecting none instead of a consensus 1120708 10_ AUS 032 Rent Rep2 genotype Markers for ke Allele Allele Allele Allele2 Allele Allele which none is T not exported in the Final Report with Valid Alleles Penta E Selecting the consensus E genotype pem The Electropherogram An electropherogram is displayed for each sample in a replicate group The electropherograms are positioned directly to the left of the report table It displays the alignment of samples within replicate groups The electropherogram field behaves exactly like its counterpart in the main analysis window draw a box from left to right to zoom in and draw a box from right to left to zoom out However allele calls cannot be edited while in the Replicate Comparison Tool Clicking on an allele call in the report table will take you directly to the marker containing that call in the electropherogram trace Double clicking on the C D or N in the status column will also display the corresponding locus in the Electropherogram 124 August 2013 Chapter 10 User Management Hi Report B 2 ij i oui cn nempe 4x 2 Rept Rep2 Alel Allelet Alele2 Allele3 Allee y 93 93 Er a p593 n t csi 0 93 n2
138. for use ofthe Software User further acknowledges that User is responsible for validating the Software for use in User s intended applications Due to the nature of computers software and installation procedures SoftGenetics cannot accept any liability or responsibility for validation of the Software in any of User s applications 3 Intellectual Property Rights The Software and any copies that you are authorized by SoftGenetics LLC to make are the intellectual property of and are owned by SoftGenetics LLC and its suppliers The structure organization and code of the Software are the valuable trade secrets and confidential information of SoftGenetics LLC and its suppliers The Software is protected by copyright including without limitation by United States Copyright Law international treaty provisions and applicable laws in the country in which it is being used You may not copy the Software except as set forth in Section 2 Software License Any copies that you are permitted to make pursuant to this Agreement must contain the same copyright and other proprietary notices that appear on or in the Software You also agree not to reverse engineer decompile disassemble or otherwise attempt to discover the source code of the Software except to the extent you may be expressly permitted to decompile under applicable law it is essential to do so in order to achieve operability of the Software with another software program and you have first requested Soft
139. from the Register Product Name Raquenin MUUTPTNEMFWROSRETARESAQURN drop down menu ee 5 SelecIf the computer License Server is being installed on has an internet connection select Online Registration If the computer does not have an internet connection or is connected to a proxy server select Offline Registration Online Registration A Locate the Account and Password on the SoftGenetics CD B Enter your Account Password and e mail address information in the appropriate fields enel Key 8 August 2013 mmm Chapter 1 Installing GeneMarker HID C The Request Code information is automatically generated by License Server D Click Register E Your software will be registered automatically A confirmation e mail will be sent to you once registration is complete NOTE Some characters can commonly be misread If you get an error qcnengestoteke affect Please make sure thatno clients are trying to register check for number 1 and lower case letter L or connected before restarting number 0 and upper case letter O confusion F Restart License Server to apply the registration information The SoftGenetics License Server Manager needs to be restarted Restart Later Offline Registration G Copy and paste the entire Request Code string and type your Account and Password information from the SoftGenetics CD into the body of an saw e mail H get iir Fieger
140. fsa 1 nie i C Forensics Frag Data NISTAPP16 PP16_MixO5case1_victim t 3 The Op en Data Files box will appear EA SIPPIE evidenoe w e TM CARA Frag Data NIST PPIB PP16 MiMO5case2_ victim ts er 4 Click Add button CV Fiag Daas APPIE PPTE MbDased ewdencelia lin ICA AF Frag Data NIST PPIB PP16 MIMO5case3_ victim t 5 The Open dialog will appear EX Frag Data NIST PPIE PP1E MIxOScased evidence ea 3 Remove Al M CS Forensics Frag Data NIST PPTB PP16_MIO5case4_victin tsa 6 Navigate to directory containing raw data files C Forensics Fiag Data NIST PP16 PP16_negative contral sa CN AForensiessFrag DataWNISTNPPTENPPT1B positive control fsa 7 Select all files by CTRL A or use CTRL and or SHIFT emis e e 3 tl keys to select individual samples Delaul 8 Click Open button in the Open dialog 9 The files selected will appear in the Data File List field T Eres m ree 10 Click OK button in the Open Data Files box and the samples will be uploaded to GeneMarker HID Features There are several features available in the Open Data Files box to make data upload easier Add Used to locate and select raw data files for upload Click the arrow button next to the Add button to see the four most recently accessed directories Remove Used to remove samples from the Data File List Highlight the sample to remove by single left clicking it in the Data File List then click Remove
141. ftGenetics project file sgf sfp NOTE Projects with similar datasets and analysis types should be chosen 5 Click Open and the second project will be uploaded to the Project Comparison tool 6 The first project originally loaded into GeneMarker HID will be marked as the Reference R gt and the second project uploaded to the Project Comparison tool is marked as the Sample S gt 7 Click the Project Comparison Settings icon to choose parameters to compare between the projects 8 Differences will be indicated in the report table on the right When a difference is selected each project s electropherogram and peak table will be displayed on the left Procedure for comparison of projects where the same samples have different names 1 Follow steps 1 4 above moe 2 Click Open and both projects will be listed at the right of 5 the screen min 3 Click on the Edit File Groups icon to group files of same Erde sample together a E 4 Click Match and Ok oo pins oe p above z MM Un a REOR xoxo un Project Comparison Tool Icons and Functions Open Project to Compare Opens a directory window for the user to identify a similar project to compare to the project already running in GeneMarker HID The first project in GeneMarker HID will be considered the Reference project and the project uploaded to the Project Comparison tool will be considered the Sample project Project Comparison Settin
142. fter alleles are edited in the Electropherogram Charts click the Update Sample Data icon in the toolbar Conflict and Suspect Marker identification will be adjusted accordingly Additionally allele changes made in the Pedigree tool will be applied in the Main Analysis window Edit Suspect Alleles Wh Pedigree Edit C Usens SoltGenetics Desktop TestPed pre i E 2 2 ib UPS Fwd 2 2 2 2 ary 2 ree M woe h 1 OSS Seach 4 Samples 6 Chats Cul M C nmtirm 8 2 LUZ Unzenfom 14 8 11 tee View History 9 enon iD 4 Perion Name Tam Sample Mie 27397930 13 Ire 95 August 2013 Chapter 8 Relationship Testing Procedure Although allele calls can be edited in the Pedigree tool it is easier to begin a relationship test analysis with good clean traces In order to begin with the best sample traces complete size calling Panel alignment and allele editing in the Main Analysis window prior to launching the Pedigree tool To launch the Pedigree tool select Applications Pedigree from the menu bar of the Main Analysis window File types accepted or generated by GeneMarker HID Pedigree module Pedigree File PRE PED Contains information about the Pedigree Tree display Individual Sample Accordance File SMP Contains Family and Individual ID information associated with specific sample
143. g Standards 1 In Size Template Editor select File New Size Standard OR click the New Size Standard icon 2 The Input Dialog box appears 3 EnteraSize Standard name and click OK 4 The Expected Size Standard frame will be blank 5 Right click at a known peak in the Sample ILS frame 7 777 a 6 Select Insert Size Du x 8 d 7 The Insert Size box appears T i ute char M rade da 8 Enter the basepair size of the peak in the Size field z 9 9 A green triangle will appear atop the peak in the Reis i Sample ILS and a new peak will appear in the ET Expected Size Standard frame diim V 3 f 10 Continue Insert Size operation for the rest of the B ee 98 0h ps H peaks in the Sample ILS Fr Tur pr 5 Panel 11 GeneMarker HID will interpolate Size values after Bos mis es two peaks are added to the Size Standard ramasa NOTE It is recommended to use the interpolated Size B n b values when creating a Size Standard due to the ec differential migration patterns of each sample Low rca 12 13 14 When the Size Standard is complete select File Save Changes OR click Save Changes icon Click OK in Size Template Editor Proceed with Run Wizard data analysis Icons and Functions The following are explanations of menu and icon options in Size Template Editor Menu Options The Size Template Editor contains three menu options File BestMatch a
144. gin analysis Run Project Opens Run Wizard for processing the data a m BEE Show Hide Toggles Displays or hides the Sample File Tree Synthetic Gel Image and Report Table frames respectively Print Report Provides the user display options for the Print Report Show Color Allows the user to select all colors to view hide all colors or choose a single dye layer Choose a single dye by single left mouse clicking on the icon m el 34 August 2013 Chapter 3 Main Analysis Overview Zoom In E EN Use the icon to zoom in on the image or hold down the left mouse button and draw a box from the top left corner to bottom right corner around the area you wish to zoom in Zoom Out Use the icon to zoom out on the image or hold down the left mouse button and draw a box from the bottom right corner to top left corner Set Axis The default setting automatically sets the Y axis according to the maximum peak intensity of the ur samples Two other options are available auto fit the Y axis using peak intensities of the alleles or the user can select the ranges for the X and Y axis Browse by All Colors Displays a comparative view of sample electropherograms by dye color Individual samples can be selected from the drop down menu Allele Call Icons These icons are only available after the raw data has been processed and the Sample File Tree Allele Call folder is selected Size Calibration Displays calibration charts f
145. gs a aa ri Launches the Project Comparison Settings box with several can options for running the comparison Marker Name Allele Name Meere Dye Score Peak Matched By Allows the user to choose the principal parameters C Matker Name Peak Size eye f n for comparison v Hegi V Comments l Dye Peak Size Alea Peak Compare Items Options for which parameters should be Peak Comparison Thieshald compared and marked as different MaxSize Dit 10 Max Height Dit 10 Peak Comparison Threshold Allows the user to qualify the ranges for detecting differences in peak attributes 120 August 2013 Chapter 9 Additional Tools Convert TXT to Binary Tools Convert Text to Binary Files The Convert Text to Binary tool allows the user to upload trace data information in Text txt file format for conversion into a four color SCF file or a five color SG1 file The SCF and SG1 files can then be read by GeneMarker HID and translated into chromatograms This tool is useful for institutions developing their own fragment analysis instruments Procedure 1 Click the Load Text File button and select Text txt files to convert 2 Once files are uploaded they will appear in the Text File field 3 The software will automatically calculate a Recommended Ratio for the user to condense the number of frames in a single trace 4 Enter a condense frames by XX number in the Condense Frames field 5 Click Export to
146. he Identity by Descent calculations the user may opt to allow for mutation allele conflicts using the analysis parameters and the AABB mutation rates Procedure Open a saved project file or import raw data and complete analysis as in chapter 3 Tools Select the desired Allele Frequncy Table or use all populations Tools Relationship Testing Tools Family Grouping Tool Group families as in the previous section Right mouse click on the alleged father node and edit node Select Alleged Father in the Contested section The PI is displayed in the table Select Calculation Details for the results at each locus Right mouse click on the table to copy paste or save as txt tab delimited file Save and Print Reports Ors7173 D2511 D75620 CcsF PO D331358 THO 013837 O16S539 0251338 D195413 D8SH73 Des 075820 CSFIPO 0351 359 THO 0138317 D158533 D281338 0195433 WA TP X D18551 AMEL Dpssme FGA PAT Ladia 1114 PAT C fa PAT 1 Fisa PAT 1 Misa PAT 2 Cha 5 Edit Individual m Individual Info Aflect Status ume 1 f Unknawn r Gerda Male Femde C Unknown cisa C hedeg Falher Corteded f Alleged Father r UHR Conbecied Ch C hes 5 From Sample Fie PAT_1_F tsa C FineiDalabare Mone z amp I Manuali Input 8 CE Activate the Save or Copy Paste for Print reports by using a right mouse click anywhere in the results table Tables are saved as tx
147. he Pedigree Tree right click the node and select Delete Node Family Name Enter a name for the family in the free form text box The Family Name field will not appear after the first individual is added to the Pedigree Tree All subsequent individuals added will be considered members of the family Person Info New Family New Individua Name Free form text box to enter a name for the individual m Display the individual s name in the Pedigree Tree by clicking the T New Individual Show Individual ID icon in the toolbar RE Gender Select either Male Female or Unknown gender for the Mox 5 Unknown individual Male nodes are squares female nodes are circles and Ta a Unaffected Gender Male C Female Unkown Unknown gender nodes are displayed as a circle within a square Father Mother When more than one mate is displayed for a Mother or Father a drop down menu allows the user to choose which 3 possible parent to associate with the child Affected Sample File 1273878 30 08 fsa gt NOTE The Gender and Father Mother options are not available when adding a mate 9 tme Affected Status Affected Status options are available to mark individual nodes for genetic linkage calculations Before marking individual nodes with Affected Status click the Pedigree Parameters icon in the toolbar and adjust settings accordingly Unknown The individual s phen
148. he Report Table and other frames in the Main Analysis window are linked are controlled by options in the View Preferences Others tab Display Settings Click the Report Settings icon in the Report Table toolbar The Allele Report Settings box will appear Select different Report Styles to see additional options After selecting Report Style options click the Save as Default icon in the bottom left corner of the Allele Report Settings box Your options will be saved and will be recalled the next time you select that Report Style Additionally select View Preferences Others Show Disabled Samples in Report to include samples that are disabled in the Sample File Tree Sort Options Sort by Marker Report Style C Allele List Forensics Bin Table Peak Table Allele Count Orientation Horizontal C Sample Name File Name Vertical Options Extend Diploid Homozygous Show Allele Name Show Size 0 1bps Show Height Show Area Show Only Uncertain Alleles iw Show Rejected Low Score Alleles Select Sort by Marker from the right click menu and choose from the fly out menu to sort Ascending or Descending If Ascending is chosen then low quality peaks will be sorted to the top of the table If Descending is chosen then the lower quality peaks will be placed at the bottom of the table This option is only available with Marker Table and Allele Count Repo
149. he dataset within a five basepair shift Ladder samples are identified by GeneMarker HID as defined in the View Preference Forensic Ladder Identifier field Major alleles and variant or virtual alleles are specified in the Control Column in the Panel editor See Chapter 5 Panel Editor This information is used for pattern recognition and automatic panel adjustment NOTE Panels that do not contain variant virtual alleles can be manually adjusted in the Panel Editor by first clicking the Major Adjustment of Panel icon then the Minor Adjustment of Panel icon NOTE Panel must be saved with signal information For optimum Auto panel adjustment See Chapter 5 Adjust Analysis Parameters After the clicking OK in the Run Wizard Additional Settings box the Data Processing box appears The raw data is being processed and sized then the filtering parameters js rmm are applied and finally a Panel is applied if selected Click OK in the Data C EM 019 59 fra Completed Processing box when analysis is complete pivot ton cometed XL 63 Conpleted Review the results in the Main Analysis window See Chapter 3 Main Analysis mss tmiss Overview If you notice many false positive peak calls you may need to adjust the x 6 suaples proce sed analysis parameters There are three options for adjusting the analysis parameters as discussed below NOTE Manual edits will
150. he position of the Bin center in basepairs Left Right Range Indicates the range of the Bin on either side of the Bin center Allele Name Peaks that appear within the Left Right Range of the Bin will be labeled with the Allele Name Control Bins marked with a 1 are considered major alleles in the Ladder sample and will be marked with an Allele Label and a darkened rectangle to mark the bin in the Main Analysis window Bins marked with a 0 are minor peaks and will not receive an Allele Label the bin will be marked with an clear rectangle in the Main Analysis window wh Help E A Qa DB i Wwe No NOTE The designation of 0 and 1 in the Control column is used to assist with pattern recognition in the AutoPanel Adjust algorithm Distance kb a Allows the user to input the distance in kb that each allele is from the beginning of the sequence For example 38 1 means that the allele is i 38 1 kb from the beginning of the sequence MM Jl I Ji NOTE This option is not applicable for forensic STR analysis E annem mpi 252 i5 laa 417 ena maai peal zz aa bq 8 67 Comments Free form text field to associate a comment with the Bin 55 August 2013 Chapter 5 Panel Editor Procedure As mentioned previously Panels are created to outline the position in basepairs of expected peaks In GeneMarker HID the Panels as
151. his option selected replicate groups will be sorted according to their order in the Main Analysis Window file tree Sort by Status With this option selected replicate pairs with Discordant calls D or Null calls N will be sorted to the top of the table Accordingly Concordant C replicate pairs will be sorted to the bottom of the Report Table Select the preferred sorting option from the dropdown and then click the refresh icon white paper with arrows to have the sorting take effect The Report Table In the replicate comparison tool synonymous replicates are compared according to the parameters set in the Replicate Comparison Settings window The results of this comparison are organized and displayed in the Report Table The replicate Group Number is in the first column of the table and the name of the first sample of each group is listed in the second column of the table under the Sample Name header Allele calls for each replicate are enumerated in the next columns and rows The Status Column The status column is the focal point of the Report Table as it displays the concordance of related replicates The status column is automatically filled with a C for Concordant if the replicates are identical or if differences are less than predetermined thresholds If the replicates are different or differences are greater than predetermined thresholds the status column is filled with a D for Discordant Finally if only one replic
152. i Lon ma rem Double click the GeneMarker HID Setup executable file EXE PFA LY NN epe ror The Installation Wizard will launch Nen The Loene Server Manager muni be natalled on ine Serve i 1 not to be ienislied cn Chere Computer Click the Next button in the Welcome window Read the SoftGenetics End User License Agreement and click the I Agree button in the Read Me File window 6 Select Install License Server Manager in the Select Program window qp zt boltieenetics Licenae Server E xd and click Next i see carm bo Soi aerate Lar Server lup pripa Tie prox am ead ited Soit meta Server on sour compules 7 Click Next in the Destination Location window Next in the Select Program Manager Group window and Next in the Start Installation REA api Lick Cancel aai and choos am zrogeami vou herve Surinam Lick h Gore seth re Sano piogiam Ther pogam i protected bu copra nos and F TV DV Ft t window to enter the LSM installation wizard 8 Click the Next button in the Welcome window 9 Read the SoftGenetics End User License Agreement and click the I Agree button in the Read Me File window 10 Click Next in the Destination Location window to install LSM in the default folder Click the Browse button to choose a different installation directory NOTE The default Destination Location for the License Server Manager program is C ProgramFiles SoftGeneti
153. i de m 99 KINSHIP ANALYSIS USING IDENTITY BY DESCENT 45552 diee oia Euh vade Soo opp ER e Ea ER uE SN ao eruta tous e ee Y pae ve oux uo vade Qe eae a 101 OVF CTI DM I RM MM am Ene eT ene 101 PROCOUUPBOS d eoa ioa marino aan EE M E ME EM N E LII EET 102 Cons COTE CEIONS escenas iai OE SOT ETE em Sansa tes TERRE MORTEM 102 DATABASE SEARCH LOCATE DUPLICATE SAMPLES AND NEAREST RELATIVES ceceecsccsccececeecsecscceceeseeeseeeceecseseeeseeeceeseees 104 OVERVIEW dux D ce MS Mudo D RAT Te ecu co e MR dE ne eer ee E 104 POCO GU Bst Oe qudm M EIER DIM ME A a Ted a 104 ICONS dio ec EDU co pd ptem act 106 Importing Population Specific Allele Frequency and Mutation Rate Information eee 108 BUNDO tae DODAS E seen nop alae LM EE 109 SAVE ana PrDE ReDOFE o ss oat eee nace e de et nba eh eae tis ER iua e e n dee E ERE IR IRR 110 AUTOMATED PEDIGREE TRIO DIAGRAMS AND ANALYSIS USING FAMILY GROUP TOOL sese 111 PEO COCO B eo do er m redu she cord ime MM E IM PI ML MM oM 111 DEGUGING Missing Parent tx E un E tee ns nea 112 Editing Personal TRITOEmatlOD constata or cm A recae E cv du bread oro aam de Pub di ads 113 DAVE ODOM cx Eco 114 PATERNITY INDEX CALCUISA TIONS 4 aie n Sa ec ila AD Cel eI 115 Sa
154. ibited under the Export Laws from receiving the Software All rights to Use the Software are granted on condition that such rights are forfeited if you fail to comply with the terms of this Agreement 10 Governing Law This Agreement will be governed by and construed in accordance with the substantive laws in force in the State of Pennsylvania United States of America June 2012 Table of Contents GeneMarker HID v 2 6 0 CHAPTER 1 INSTALLING GENEMARKER HID EEANEYEER RES SFO PAN FAS QUAE E RATAT EFE Se pA T ERE CE FR EATER 4 COMPUTER Had TU EPIS SEL O n ids t UN Eee Een 5 VALIDATION VERSION M T 5 DIDS MUA TTE EE 5 VOCAL LIGENSING OPTION m 6 PASO OUI ERROR T 6 TRO SOO c 6 B foro 7 IE TW 7 Install License Server MANAGED scccuncscccnnesecsseeccsaseessaeeeesaueeeesausessauueessauseeesausesssauseesausesssaansesauascssauesssaaages 8 Register License Server Manager for GeneMarker HID Usage eise nnne nnne nna 8 Install GeneMarker HID Software on the Cli
155. ile in order to import additional genotypes 8 Select Ok in both the Genotype Editor and New Individual Dialogs to display the node of the selected individual then right click and select find family as in the above searches to search the database a Retstonship Testing New File i mur file Datalave Tool J TEH Br TS S UNT Make Samet 111 Renon By E E hrad ndn uid MISTE ru WY iird Sl E wy XO Tert Tan DEAL 27 Sampl T5 JEET latar Ton p05117 DEISI pesnen CSFIFU D33139 THil D135317 En ps4 bil 338 105433 vivi TRO AMEL CESAIE FEA Mother sagin T Ti Hasia 105 August 2013 Chapter 8 Relationship Testing Icons and Functions a Relationship Testing Relationship Testing Main Drop down menus Detainee Tool Select from File DataBase or Tool options Relationship Testing Tools include n Family Group Tool for automated pedigree trio drawing If Relationship Testing Allele Frequency for major US populations File DataBase Tools Mutation Rate specified by AABB i re E E Family Group Tool Population Statistics for the file under analysis Allele Frequency Genetic Analysis Settings allows setting the above options at the luris Foie same time additional settings include limiting the number of files retrieved in the database search by a minimum number minimum
156. ill be highlighted green in the File Name List Match by Fixed Position Allows the user to manually identify the characters of the filename for grouping the samples Section Separators like are counted as individual characters Group Identification Enter the number of the beginning and ending character to identify how to group the samples The section of the filename specified will be highlighted red in the File Name List Control Identification Enter the number of the beginning and ending character to identify which part of the filename contains the control identifier The section of the filename specified will be highlighted green in the File Name List BH File Name Group Editor om w on w m www i 1 1 1 i 1 41 1 SH Cow UG ow io HM Mon HM P 38 eee n ow amp Control Identifier Enter the character from the Control Identification section highlighted green that describes the control or reference sample Example N normal or R reference Select Case Sensitive if the Control Identifier needs to be identified by upper or lower case letters Control Match Mode Choose either Whole Words or Include Whole Words should be used if the characters entered into the Control Identifier field need to match exactly Include should be selected if the characters in the Control Identifier field only need to be identified in the filename i e not an e
157. in Table NOTE Allele Count requires that a Panel is applied to the data See C Peak Table Chapter 5 Panel Editor Heje Count Fe atures Sample Name File Name Orientation Orientation xL 5 Or Horizontal Sample names appear on the left in rows and Markers Horizontal C Vertical Show Rejected Low Score Alleles appear at the top in columns IV Hide Extra Sample Names Cancel 70 August 2013 Chapter 6 Reports and Printing No Samples 185386 pressos remar p2151411 Total Numbg Vertical Markers appear on the left in rows and 22 sample names appear at the top in columns 2 osz_aca scr a 052 04 55 SCF l s josz mos scr ofl0W_ixejected LOW _ 5COTE ZXLLCLES 7 Show Rejected Low Score Alleles When selected the is Joer soseer peaks with peak scores below the Run Wizard e oez cos scr 7 063 D05 SCF Additional Settings Allele Evaluation Peak Score Reject RET setting will be displayed in the table 5 oes ros scr Hide Extra Sample Names This feature is not active for Allele Count Report Style NM MM MM RN ND Oo ND PIN NI NM NT NM PN N N Oj Fr Fr er Ni RP RP NM NM N eR Ri a N ND AD o NM NM HM HM HM HM FP NM HM NM NM 1 Print Report The GeneMarker HID Print Report displays Electropherogram and or Peak Table information for all or selected samples in a dataset To access the Print Report go to Project Print
158. in rale Lando ICT IE PAT 4 Bicis M ja Hl i i ss pe tC fae 1 6 15 GeneMarker HID will appear Navigate to the Burst B r4 5n 19 directory containing the SGF or SFP file and click dans s S B rir EMI Open Additionally the last four projects that were pay TE opened by GeneMarker HID appear when the File gt s ug s 1 8 ug zo 23 a 15 Eese Up Page Down 74 August 2013 Chapter 7 Mixture Analysis 75 August 2013 Chapter 7 Mixture Analysis Chapter 7 Mixture Analysis Chapter 7 Mixture Analysis Overview Procedure Icons and Functions What to Expect Save and Export Results Search Database Mixture Analysis Equations 76 August 2013 Chapter 7 Mixture Analysis Overview Mixture analysis is required for many samples that are not single source as in the case of some crime scene and missing persons and unidentified human remains applications The mixture analysis application was developed using recommendations of the DNA commission of the International Society of Forensic Genetics Gill et al 2006 and methods of Clayton et al 1998 and Gill et al 1998 GeneMarker HID mixture analysis follows the steps involved in the interpretation of STR mixture data Clayton et al 1998 Identify the Presence of a Mixture Designate Allele Peaks Identify the number of potential contributors Estimate the relative ratio of the individ
159. in the laser path Spikes are typically less than a base pair wide Do not select Spike Removal when 214 Derivative Trace has been applied Size Call GeneMarker HID offers two sizing methods Local Southern Used in most genotyping software applications and is recommended for most analyses This method is based on the idea that smaller size fragments run faster Plot a size v time graph and overlay a size v 1 time graph to determine linear trace Southern E M Measurement of DNA Length by Gel Electrophoresis 1979 Analytical Biochemistry 100 319 323 Cubic Spline Method Cubic Spline is offered as an alternative method that may be more appropriate for some data This method uses a cubic equation to connect known points on the size v time graph An example of a cubic equation ax bx2 cxt d The Astrophysical Journal December 1 1994 436 pages 787 794 Local Southern Method Cubic Spline Method Three sections used in cubic equation Region of Interest 17 August 2013 Chapter 2 General Procedure Allele Call The Allele Call section allows the user to set allele calling range detection thresholds and filters Auto Range The software will identify peaks in the processable data range for each lane Manual Range To select a specific analysis region de select Auto Range and input the desired base pair range Peaks outside the Manual Range will not be called Peak Detection Threshold NOTE The Peak Detection Thre
160. ine Subtraction Use 2076 of lowest intensities to the right of the beginning of the range Looks at trace in 500 600 frame sections Chapter 2 General Procedure Pullup Correction Ax B A being the major coefficient Input matrix or use single dye adjustment up to 0 20 for small corrections When Manual Pullup correction is chosen a txt or mtx matrix file can be uploaded and used to deconvolute dye colors NOTE De select automatic Pullup Correction in the Run Wizard Data Process box if a manual matrix correction has been applied Saturated Peak Correction ABI instrument saturated peaks are typically 28000 RFU The top of a saturated peak looks split A small pullup peak may be present under the saturated peak GeneMarker HID takes the small pullup peak and adds it to the split in the saturated peak Spike Removal Caused by overheating of camera chip voltage spike etc Spikes usually only 1 2 frames wide peaks usually 5 10 frames wide Create a first derivative trace of the raw data Spikes are the 1st DT outliers 3 5 sigma 14 August 2013 Chapter 2 General Procedure Second Derivative Trace A1 A2 A2 A3 A1 A3 2 A2 Use when you have a fat base to your peaks ex Dye blob under peak etc NOTE Do not use 274 DT with Spike Removal because real peaks look like spikes Process Data After the raw data files have been uploaded to GeneMarker HID they are ready to be processed The processing step includ
161. ing GeneMarker HID Local licensing Option GeneMarker HID v2 0 and above supports text based registration for the local licensing option no USB device dongle key or hardware is required This text based registration ID is registered to one specific PC If the license needs to be transferred to a different PC registration for that one license PC must be inactivated first before the software will be registered to the new PC Installation 1 DIG qe Insert the SoftGenetics CD into the optical or CD ROM drive If your computer is not set to automatically open a CD navigate to the optical or CD ROM drive on the computer and open the directory Double click the GeneMarker HID Setup executable file EXE The Installation Wizard will launch Click the Next button in the Welcome window Read the SoftGenetics End User License Agreement and click the I Agree button and Next in the Read Me File window Select Install GeneMarker HID Recommended in the Select Program window and click Next Click Next in the Destination Location window to install GeneMarker HID in the default folder Click the Browse button to choose a different installation directory NOTE The default Destination Location for the GeneMarker HID program is C ProgramFiles SoftGenetics GeneMarker HID ver 8 Click Next in the Select Program Manager Group window to accept the default Program Manager Group NOTE Changing the Program Manager Group default may affect p
162. is followed the specified ladder electropherogram will be fixed at the top of the page and the page down key will open the sample electropherograms allowing easy comparison of the allelic ladder and each sample 64 August 2013 Chapter 6 Reports and Printing 65 August 2013 Chapter 6 Reports and Printing Chapter 6 Reports and Printing Chapter 6 Reports and Printing Report Table Print Report CODIS Report Save Project 66 August 2013 Chapter 6 Reports and Printing Report Table The general features of the Report Table were outlined in Chapter 3 Main Analysis Overview Here we will discuss and give examples of each Report Style available in the Report Table Allele List The Allele List Report Style displays the basepair size or Allele Label if a Panel is applied of the called peaks The sample names are listed in rows in the far left column and peaks are numbered in columns at the top of the table EET Ron Sti Features ends Report sample by Sample Name or File Name Bin Table Show Only Uncertain Alleles Bie ods When selected displays only the peaks with Quality ranks of Check yellow a and Undetermined red C Since Hane 1 Fie Mime Show Rejected Low Score Alleles nd te When selected the peaks with peak scores below the Run Wizard Additional FF Heb Eus Sarl Hanse Settings Allele Evaluation Peak Score Reject setting will be displayed in the table EM UN ee ee XI 3737373 Fo a AE go F o
163. isplays a comparative view of sample M MN I Marker PAT 1 Cha mj PATILF he FE Sbr ERI Hal Sibe LR eoa LA electropherograms by dye color Individual samples can be el m m EXEC OM selected from the drop down menu wm ir A voll LE Cacao Db165533 16 3 10 12 1557273 135636 LEE 131818 bray i2 13 8 3 1319790 acs deny Meme Save Kinship Report as txt file or right click to copy paste Ta AR fL cm uer b tape aT to anti cae fan 18551 8 15 15 16 1 49870 usss 1 24565 112453 p21511 8 23 a 11 15225 083113 108112 on UENIT ORAT ns Database Search Locate Duplicate Samples and Nearest Relatives Overview Missing person identification may be necessary in many situations mass natural disasters earthquakes tsunamis human attacks Sept 11 World Trade Center war or in the cases of thousands of missing children and adults reported every year The database function of GM HID is capable of closed system searches for same sample matches in the case where a DNA sample of the missing individual is available from personal use items In the case where there is no personal sample available the kinship formulas IBD are used to find highest likelihood ratio scores for each relationship level The database includes all CODIS markers and should be used with complete profile samples The Save to DataBase function allows easy updates of the relationship testing database It is strongly recommended to b
164. izes of standards used for size calling Peak Position in frames and Size sizes are automatically entered but easily edited 47 August 2013 Chapter 4 Fragment Sizing Standards What to Expect It is important to verify sizing accuracy prior to analyzing a dataset If a sample is not sized correctly peaks may be called Off Ladder OL if a panel is applied Incorrect sizing most dramatically affects larger size fragments Before amp After Editing Size Call t ap zl DRE LL 22 65 N sa BELAT PES Jul ee ee ee STITT EENET amos JANGNI 200025250 TMS INITIO 5p 500 1 CHR OR CSET qus 48 August 2013 Chapter 5 Panel Editor 49 August 2013 Chapter 5 Panel Editor Chapter 5 Panel Editor Chapter 5 Panel Editor Overview Procedure Icons and Functions What to Expect 50 August 2013 Chapter 5 Panel Editor Overview The Panel Editor can be accessed from the Tools menu in the Main Analysis window OR via the Panel Editor icon in the Run Wizard Template Selection box The purpose of a Panel is to outline the position of expected alleles Loci or Markers give a range where a group of alleles is expected to appear and Bins indicate the specific basepair position of the expected allele In GeneMarker HID Markers are indicated by a horizontal gray bar across the top of the electropherogram Bins are indicat
165. k computer Exit Closes the Panel Editor tool Be sure to save changes to the Panel before exiting Tools Menu Match Ladder Opens the Select Ladder box Choose an allelic ladder sample from the drop down menu Click OK and the Panel will adjust slightly to align with the peaks in the selected ladder sample NOTE Large differences between peak and Bin position cannot be resolved with the Match Ladder function Virtual Panel Select a Panel from the Panel List and click Virtual Panel in the Tools menu The Create Virtual Panel process box will appear Click OK and a new Panel will be added to the Panel List labeled VPanel_PanelName This newly created Panel is an adjusted version of the original panel selected in the Panel List GeneMarker HID attempts to align the original Panel to the Ladder sample peaks based on an average calculation NOTE It is recommended to use Virtual Panel only when small adjustments to the Marker and Bin placement are required Use Major Panel Adjustment icon for larger adjustments arm decr Acton be Export the Project Panel mm Exports the currently selected Panel in the Panel List as an XML file to a specified directory on a local or network computer EP A Draw a box from left to night while holding the le amp mouse Scroll Graphics Vel right wtale holding the nlt mouse Help Menu The Help menu contains a link to Hot Keys in Panel Editor Click Hot Keys and the 7 7
166. k Table Delete Undelete Allele Right click at the vertical grey bar indicating the center of the called peak or the peak cell in the Peak Table and select Delete Hot Key DEL To call the allele again right click the peak and select Undelete Hot Key SHIFT DEL Confirm Unconfirm Allele Edit Allele 33 Marker Penta D Allee 182 Size 181 7 lw Confirm the Allele Cancel If a peak is given a low quality score it will receive a Check yellow or Undetermined red Quality ranking To give the peak a Pass green Quality ranking right click the peak center bar and select Confirm Hot Key 26 August 2013 Chapter 3 Main Analysis Overview CTRL M The peak will be marked Pass green and receive a Confirmed comment in the Peak Table To un confirm the allele select Unconfirm from the right click menu Hot Key CTRL ALT M Confirm Unconfirm All Confirm All and Unconfirm All options perform the same actions as the Confirm Unconfirm allele except that the Quality ranking for all peaks in that dye color for that sample will be affected Edit Allele Right click an allele in the Electropherogram or Peak Table and select Edit Allele The Edit Allele box appears Add or change the values in the Allele and or Size field The Allele field will be blank if no Panel has been applied to the dataset Check Confirm the Allele to automatically give the peak a Quality rank of Pass green
167. k Table respectively Bw 1 ne 9 Di T tn Erw 1377 2716 097 p35 Costum Start Area Ratio SM OM mS wal ce a eu 1650 acon Al r t The value obtained when the peak s area is divided by the EE n f Serou conos area of the highest peak in the dye color or Marker Bs 153 zn nu wal Bo feme 23 zn 3 MWh 18531 2 8 053 View Festony po 3 2227 D AMA Marker Panel Only Indicates which Marker Locus the peak is contained in Allele Panel Only Indicates which Bin the peak is contained in Difference Panel Only Indicates the absolute value in basepairs of how far the peak center is from the Bin center Quality Panel Only Assigns a Pass Check Undetermined quality ranking for each peak with regard to the peak Score as set in the Run Wizard Additional Settings box See Chapter 2 General Procedure and or software editing of the original raw data such as correction of saturated peaks SAT Repaired Score The peak quality score is calculated based on signal to noise ratio and peak shape or morphology Lower scores indicate poorer quality peaks Additionally the Score value is a based on an exponential curve Start End Indicate the beginning and end of the Area calculation for the peak Allele Comments Software and user edited comments appear in the Comments column Sample Comments Added by the user with a right mouse click on the sample name in the sample list
168. k intensity ie en values for all positions including Negative positions k Bk Cancel Show Peak Area Displays the peak area value for all Positive and Suspected peak positions Dollar signs separate values if more than one display option is selected Orientation TR EL Horizontal Sample names appear on the left in SOD REES QQRE 088 wef za rows and Markers appear at the top in columns Report GE Bin 3 Hev fraa 001 HOT fsa x gt Sy rer Vertical Sample names appear in the far left column in rows Markers and Alleles in order of 2600 4 basepair size appear at the top in columns Show Only Uncertain Alleles When selected displays only the peaks with Quality ranks of Check yellow and Undetermined red Show Rejected Low Score Alleles When selected the peaks with peak scores below the Run Wizard Additional Settings Allele Evaluation Peak Score Reject setting will be displayed in the table Hide Extra Sample Names When data is displayed in Vertical Orientation the sample names are repeated for each row of data that the sample is associated with If Hide Extra Sample Names is selected then the sample name will only appear l dun nent puer S Ries once in the first of the rows it is associated with 68 August 2013 Chapter 6 Reports and Printing Additional Functions Allele Editing Options
169. k symbol next to the individual s node Suspect Markers include those which contain additional peaks or low quality peaks Mouse over the node with suspect Markers and a list of Markers or loci with suspect peaks will appear Single left click the Marker name and the individual s electropherogram will appear in the Charts tab on the right The suspect Marker will appear red 94 August 2013 Chapter 8 Relationship Testing Show Inheritance Conflicts JE Peeper Fart ew File TE B reece e omen Sese x amps MAS Chor 27907810 Dia e E 35 a nsa z 02 9 oes 3 2030 Peso Person Kammer Fam Tampe l ile 275875730 11 a Samples List The Samples List displays the filename and individual ID for each sample in the current dataset Drag and drop samples from the Sample List to nodes in the Pedigree Tree to associate individuals with the family Electropherogram Charts The electropherogram traces for the samples in the Sample List appear in the Charts tab Double click a node or single click the Marker name in the Conflict or Suspect list of the Pedigree Tree and the electropherogram trace for the Marker will appear If a Child node is selected the Mother and Father traces will also appear in the Charts tab Navigation and allele editing in the Electropherogram Charts is similar to navigation options in the Main Analysis window See Chapter 3 Main Analysis Overview NOTE A
170. k within the marker Stutter Filter Forward and reverse stutter peaks commonly caused by PCR chemical reactions can be removed using the Stutter Filter In forensic analysis the stutter positions of interest are N 4 N 8 and N44 relative to the highest peaks in the marker given tetra nucleotide repeat units The x refers to the number of nucleotide repeats specified in the Marker Parameters Nucleotide Repeats field The settings are in percentage of the primary peak Peaks in the N x N 2x N x positions that are below the stutter threshold will not be called NOTE The stutter filter is meant to eliminate the extra steps of analyzing peaks caused by instrument anomalies Set all Stutter Filter settings to 076 if all peaks must be called Apply Filter to All Markers When selected the values in the filtering parameters fields will be applied to all Markers in the Panel To optimize the stutter filter set each marker with stutter values determined during your lab validation procedures Edit Marker Bins Right click the Marker bar and select Update Alleles The Update Marker Alleles box will appear Adjust parameters and click OK Nucleotide Repeat Auto Detect Based on the peaks present in the Overlay View GeneMarker HID will attempt to detect the number of nucleotides in each repeat unit of the alleles and place Bins at the appropriate interval Set by Manual Select this option if the number of nucleotides in the allele repeat u
171. label and the peak table below the electropherogram in the quality reasons column Right click the sample filename in the Raw Data or Allele Call folder to see additional options Sorting Options Select Page Opens electropherogram traces for the number of samples specified in the View Preference Display Settings Max Chart In Page field Hot Key Page Up Page Down Select Next Group In descending order selects the same number of samples previously selected by Select Page grouping options see Sample Grouping section below or double click option Select Max Opens electropherogram traces for the number of samples specified in the View Preference Display Settings Max tt Open Charts Fist Orde FieNam Deselect All Second Order Well ID v Unselects all selected samples in the Sample File Tree list and closes the Third Dider Size Score electropherogram traces z PB iW Ascend Sort iw Disabled to Bottom Sort Samples Case Sensitive vw Ordered by Group Opens the Sort Sample Options box Select First Second and Third Order E ene ae NA sorting from the drop down menu options Sample Type File Name Lane m Cancel Number Well ID and Size Score Hot Key F3 Search Options Search File Opens the File Search box Enter any part of a filename to search for the sample in the list Click the Search button Left click and use CTRL or SHIFT key to highligh
172. layed in the electropherogram default is to display allele labels of EERE dye 1 only when all dyes are overlaid in the electropherogram Lira e Pe 1 Chart Settings T es a Max of Open Charts Select the maximum number of samples you would like to display as an electropherogram at one time Max 96 Hi cerei Use the Sample File Tree right click option Select Max to open the 29 August 2013 Chapter 3 Main Analysis Overview number of samples specified Max Chart in Page Select the maximum number of sample electropherograms you would like displayed in the Main Analysis window at one time Max 8 Use the Sample File Tree PageUp Down option to select subsequent groups of samples Max Allele Label Layers Select the number of allele label layers to view at once Max 10 This determines how far you must zoom in to clearly read neighboring allele labels and affects how the print report will be displayed Show Loci with multiline Select this option to display the names of all markers above the electropherogram when two or more dyes are displayed Show Saturation Alert Line Displays a red horizontal line under the X axis in regions where saturation and or elevated baseline were detected in the raw data Peak Label Choose up to four labels size height area score to display as a flag next to individual peaks in the electropherogram Position Choose to place the peak label at either the top of
173. le Sample ILS The Sample ILS frame displays the selected sample s ILS trace Click the Show Dye icon in the toolbar to cycle through the other dye colors Right click at a peak without a green triangle indicator and choose Insert Size The Edit Size box will appear Adjust as necessary and click OK The green triangle will now appear atop the peak and also in the Expected Size Standard Match Score Appears in the upper right corner of the Sample ILS and corresponds to the degree of pattern match between the sample s ILS and the Size Standard selected Perfect matches receive a score of 100 no correlation receives a score of 0 Navigation in the Sample ILS frame is similar to the navigation options in the Main Analysis window See Chapter 3 Main Analysis Overview Procedure As mentioned previously Size Standards are created to assign basepair size information to fragment peaks in a sample ILS The other dye color fragment peak positions are then interpolated based on a linear size scale from the basepair sizes assigned to the peaks in the ILS GeneMarker HID s Size Template Editor tool allows users to apply pre defined commercial Size Standards or create new custom Size Standards based on the dataset ILSs Pre defined Size Standards include 5C120 GS500 ET400 R GS500_1 ET550 R HD400 ET900 R ILS500 GS 100 250 ILS600 GS 75 300 Liz120 GS200 Rox1000 GS350 SEQ 600 GS400 Pre Defined Size Standards There are two ways to choose a pre defin
174. lick Register E Your software will be registered automatically A confirmation e mail will be sent to you once registration is complete NOTE Some characters can commonly be misread If you get an error trying to register check for number 1 and lower case letter L or number 0 and upper case letter O confusion F Launch GeneMarker HID and begin analysis Lom ad Teed Key Offline Registration f rege Dine c Pags ia A Copy and paste the entire Request Code string and type your Account and Fesescem Password information from the SoftGenetics CD into the body of an e mail Pede CO O 5 3 B Send the email to tech_support softgenetics com C The Registration ID will be sent to you via email within one business day ee D Copy and paste the Registration ID from the e mail into the Registration ID field oec nose E Click Register F Launch GeneMarker HID and begin analysis Upgrade Installing Over the Previous Version If you choose to install the new version of GeneMarker HID over the previous version you will need to choose the same directory for installation Several of the old files will be replaced with newer files Other files that are not present during installation but are created during analyses or by the user will remain in the folder and can easily be recognized by the new version of GeneMarker HID Please make a backup copy of the targ
175. likelihood ratio gender known or gender not known and Advanced settings to limit the search to specified relationship levels the default is to search and display potential same individual father son mother daughter sibling and half sibling Population Statistics Kinship Analysis Genetics Analysis Settings Select an Allele Frequency Set E 215 xil Allele Frequency Tables Allele frequency tables for major US populations may be selected nm Hen DESO w e 6 5 gi from the drop down menus in the Select Allele Frequency a los ta m mi x te Settings box If results of all populations are preferred select the Wee tae ee Use all Populations box and the final report will append with the uum HIVE RUP gus Gs results using each of the tables sequentially The Delete button _ ee 1 Bem may be used by individuals with access rights to remove any TE CT AC RO MT 3 ia population frequency tables that are not needed by the 5E A i Ho laboratory Use the Open folder icon to import formatted allele xam re frequency tables for other populations Allele Frequency Tables and Sources PowerPlex 16 2001 Levedakou et al Caucasian American African American Hispanic American and Asian American Levedakou E N et al 2001 Allele frequencies for fourteen STR loci of the PowerPlex 1 1 and 2 1 multiplex systems and Penta D locus in caucasians african am
176. lity of the Software in accordance with the Documentation Permitted Number means one 1 unless otherwise indicated under a valid license e g volume license granted by SoftGenetics LLC Computer means an electronic device that accepts information in digital or similar form and manipulates it for a specific result based on a sequence of instructions SoftGenetics LLC means SoftGenetics LLC State College PA 16803 2 Software License As long as you comply with the terms of this End User License Agreement the Agreement and pay all license fees for the Software SoftGenetics LLC grants to you a non exclusive license to Use the Software for the purposes described in the Documentation Some third party materials included in the Software may be subject to other terms and conditions which are typically found in a Read Me file located near such materials 2 1 General Use You may install and Use a copy of the Software on your compatible computer use the Software on a computer file server provided concurrent use does not exceed the Permitted Number No other network use is permitted including but not limited to using the Software either directly or through commands data or instructions from or to a computer not part of your internal network for internet or web hosting services or by any user not licensed to use this copy of the Software through a valid license from SoftGenetics LLC and 2 2 Backup Copy You may make one backup copy of the
177. llele Labels appear below the electropherogram Label Peak Height Ratio Chart Height mm B Abide by Panel Prints only alleles within a Panel Alleles that are outside the Amm m Panel are not included in the printed report New Page for Each Sample Prints a new page for each sample instead of eem aw toe continuing on the same page as the previous sample not available if Grouped by Dye is selected Auto Scale Markers When selected the RFU intensities of low peaks are adjusted to match the intensity of the highest peak in the dye color When low peaks are increased the intensity magnification factor is noted in the Marker 2X 8X Grouped by Dye Organizes the electropherograms in the Print Report such that samples are listed in order of dye color selected i e all samples in blue first then all samples in green etc often used in combination with Print Samples with Grouping option 72 August 2013 Chapter 6 Reports and Printing Mark Deleted Edited Peaks Prints an x above a deleted peak and an E above an edited peak in the electropherogram Label Peak Ratio Select this option to print the peak ratios on the electropherogram of the print report available when the View Preferences Display Settings is selected for peak top Chart Height Use this feature to customize the size of the electropherogram in the print report Print Samples with Grouping print and or save samples by group when the grouping
178. lor in the Print Report Mix Dyes Prints all selected dye colors on one electropherogram Hide Bins uncheck this box to remove the bin markers from the X axis of electropherograms in the print report Advanced Options Print Report LI Print Project Comments Includes the Project Comments at the top of the Print Pint Type Report Select Each Page option to display the Project Comments on each page Pei ERE in the report Saner Dyes All Samples VV Dye VW ut 1 f Selected Samples jv Dve2 v Dyeb Print Report Header Includes Institution and User ID from User Ae ike Management and Template Panel Name from the Analysis Template 7 Electophetogram 2 Died PeakTable Label Dyes amp Peak Numbers Labels dye color with number of peaks for each Foll M m electropherogram Hide Bins Forensics Table Implement Y Axis Settings Prints the report using the Y axis settings the user selected in the Main Analysis window Set Axis icon Shas Tour iw Print Project Comments on Each Page Word Wrap Chart Height mm Specify the size of the printed electropherograms Wi ues Label Dyes amp Peak Numbers Iv Print Markers Minimum 10mm maximum 100mm Implement Y Axis Settings iw Print Alleles gt 5 Abide By Panel Print Markers The Marker label bars appear above the electropherogram SEREA ETR Mark Deleted Edited Peaks Label Peak Ratio Print Alleles The A
179. low the Run Wizard Additional Settings Allele Evaluation Peak Score Reject setting will be displayed in the table Hide Extra Sample Names When data is displayed in Vertical Orientation the sample names are repeated for each row of data that the sample is associated with If Hide Extra Sample Names is selected then the sample name will only appear once in the first of the rows it is associated with Bin Table If a peak is detected in at least one sample the Bin Table Report Style will report the presence or absence of a peak at that position for the rest of the samples in the dataset Features Allele Report Settings mm Options Report Style Options Abide By Panel When selected the table will show only called ine eh V Abide By Panel alleles within Panel Marker ranges This option is only active when pri E oae a Panel is applied to the data Bin Table Hose Show Type Symbol Enter values to indicate the presence of a peak at Suspected P the position Positive the absence of a peak at the position bah es Show letensiyy 7 Negative and a Check or Undetermined Quality rank at the Somple Name FieName V Pon Pei Area position Suspected emir Show Intensity Raw Displays the peak intensity RFU value for all Merian Show Only Uncertain Alleles Positive and Suspected peak positions A 0 value is given to t Had lt VETE J Show Fejected Low Score Alleles Negative positions Selecting Raw will show the pea
180. m out and set the axis ranges respectively These icons are synonymous with their counterparts in the Main Analysis Screen Browse by All Color E Opens the All Color Browser which allows the user to see the alignment of each separate dye trace for a given sample Use the sample dropdown menu in the upper right corner to change samples Show Chart Table Displays a chart below each electropherogram which includes information such as peak size heights and user comments Right click on the table to modify its contents Save Report Use this icon to save the Report Table as a tab delimited text file Click the inverted triangle to switch between export formats All exports contain a header with project and analysis information Whole Report With this option selected the Report Table will be exported in its entirety Final Report Exports a header the status column and the final genotype column only Final Report With Only Valid Alleles Identical to the Final Report option with the exception that markers with none selected as the final genotype see below will be excluded from the export Regardless of which option is selected the user will receive a warning if they attempt to export the report table if Discordant D calls are present Use the pop up dialog box to bypass this message Sort Report s Use this icon to sort replicate groups in the Report Table There are two sorting options Sort by Group Sequence With t
181. ment or report Or export as a txt file DatsBase Tools File jesu TAE Fr Qe di Save As Name Date modit Type i dk PAT data Recert Places E Sasi B new RT CO ER Size l Famdy Fem Make AF Markers xA Samples WA Chats B Report Calculation Deiat _ Pie kae Same Penn 1 1 678 XY Fate PAT 1 Fh Mowe PAT_1_M laa sa MA 615 26 Ful S s PAT 1 C h Ta 23 2 miia na 23 26 Tha 2 PAT 3 Fha PAT 5 Clas LEES EELE 7 724 PAT 5 F ha 740 PAT amp Clea 710 10 26 118 PAT 3 Clea 55A 9 26 Hob Ste 110 August 2013 T Sech AR Copy Tatle Export Report VEN 207E 13 207E 13 T 24E 04 Chapter 8 Relationship Testing Automated Pedigree Trio Diagrams and Analysis using Family Group Tool When a naming convention is followed the Family Group Tool enables matching of files into family groups Mg File Name Group Editor Fite Name List MaohedGioups E 1 T2 3 PAT_ _F fea 2 04 Kr ess 2 2 Cl PAT_2 F fea Procedure ce T REX tse 4 FAT 4 m 5 5 PAT 5 F iss 6 PAT 6 Fisa F IFAT 7 Cha 7 Fiza 5 PAT Ladder iie PAT Ladder 2119 1 Select Applications Relationship Testing from the menu bar of the Mai
182. menu allows the user to create save and export Panels The Tools menu contains options for datasets with allelic ladder samples and exporting a Panel The Help menu contains navigation hints for Panel Editor File Menu Create New Panel Launches the Create New Panel dialog box with the options to create a new Panel Automatically or Manually Delete Current Panel Marker Deletes the Panel or Marker that is currently highlighted in the Panel List Save Changes Saves edits and changes to the Panel in the SoftGenetics GeneMarker HID Panel directory Hot Key CTRL S Save as New Panel 58 August 2013 Chapter 5 Panel Editor Opens the Input Dialog box with a field to enter a new Panel name The Panel is added to the Panel List and saved in the SoftGenetics GeneMarker HID Panel directory Import Panels Opens a Windows Explorer window to the same folder the sample files were uploaded from Use the Import Panels option to find previously exported Panel Files xml on local or networked computers Import Pre Defined Panels Opens the SoftGenetics GeneMarker HID MLPA Panel folder Use this option to find additional MRC Holland MLPA Panels Import ABI Panels Launches the Import Panels from GeneMapper box Opens Panels and Bins Text files and converts them to single Panel files in XML format for use in GeneMarker HID Export Panel Exports the currently selected Panel in the Panel List as an XML file to a specified directory on a local or networ
183. mily Full Sibs Una ERA Shee Mukslien Halin AT Edit Node Half Sibs 1 PAT_13_M fsa x 4 51E 00 Farmal and khabar bidaian Liffenerer Add Child 2 PAT_2_M fsa MX 4 06E 00 3 PAT 5 C sa XX 2 32E 00 Add Mate 4 PAT 12 C fsa XX 2 00E 00 Pew Posey A 5 PAT_5 F fsa xY 1 96E 00 Find Family B PAT_ _Cfsa XY 1 67E 00 Snach Meret Displuy 7 PAT 12 M fsa XX 1 49E 00 ez Find Individual 8 PAT 3 F sa xY 1 44E 00 F Benen Drinin Biria LA 10 000 Maus Taa Humber 10 Delete Node BS Diniy Geroin he Dupi Locus eerie Export Bitmap Save Paama when Save Hai Sead 5c pir B b Fahan BE Wolha Taug 104 Fubsie Habib August 2013 E teen Chapter 8 Relationship Testing f e Cota Took Sample in Pedigree Tree 1 Import or draw Pedigree Tree See Chapter 7 Pedigree Analysis and Automated Pedigree Tree 2 Right click on the node and select find family from the drop down menu 3 Left click on the Report icon in the tool bar to display the file name and any duplicates of that file found in the data base The samples with the highest LR for each relationship type are displayed in descending order in addition to the sex number of matched alleles and matched markers Search Database for txt file genotype 28 TH BF FAA Fare RT ate fainter sase an ey Saga A a Poet Caiculaen Dota te hore a averlo ML Cs Fates a
184. n Analysis window 2 Match by Sections Positions or Group Order and then Match Whole Words OK 3 The Pedigree for the families is drawn and displayed 7 at the left of the Sample List bi ca Compete Character Control heed ecd ion Maleh io lderttie 75 T Ji Relationship Testing 4 Right click on a node for edit or omsme J w ud BT FAA rwv Maier AE Makers z sese analysis options Semis U Crate B mer B Caci Da 5 Select Family displays all electropherograms for the pedigree ni tree at the right generos 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 279 6 Select Node or parents siblings Va displays electropherograms anon x 5 7 Edit Node allows editing of file or ais ere SR EE ER EE EEE electropherogram information Be ee sure to use the refresh key after any Finda changes en ec 8 Add Mate or child to expand the mete pedigree cia Testing Fie Dwtw sse Tools es iTH BIT T Alia Actas uA Waker 5 055008 se ac D Sample UL Cheer Aecot Detak BAGE DES 72 14 12 15 BL posit 29 3 58 8 Tat pero 1 now 151156 1 8 14 15 6 Trot 93 33 93 931 D13531 14 11 12 3 Diss M 13 n Allele conflicts are listed and the node is sss moo 14 VA 13 1 14 14 highlighte
185. nd Help The File menu allows the user to create save and export Size Standards The BestMatch menu contains options for selecting a Size Standard The Help menu shows navigation hints for Size Template Editor File Menu New Size Standard Opens the Input Dialog box with a field to enter a new Size Standard name Follow the steps above Custom Size Standard Creation Delete Current Size Standard Deletes the Size Standard that is currently highlighted in the Size Standard List NOTE This action is irreversible Save Changes Saves edits and changes to the Size Standard in the SoftGenetics GeneMarker HID Size Standard directory Save as New Size Standard Opens the Input Dialog box with a field to enter a new Size Standard name The Size Standard is added to the Size Standard List and saved in the SoftGenetics GeneMarker HID Size Standard directory Import Size Standard Opens a Windows Explorer window to the SoftGenetics GeneMarker HID Size Standard directory Use the Import Size Standard option to find previously exported Size Standard Files xml on local or networked computers Export Size Standard Exports the currently selected Size Standard in the Size Standard List as an XML file to a specified directory on a local or network computer Import ABI Size Standard Opens a Windows Explorer window to the same folder the sample files were uploaded from Export ABI Size Standard 42 August 2013 Chapter 4 Fragment Sizing Standards
186. nd limits based on LR or minimum number of retrieved samples Show Conflict Toggle between Show Conflict with Parents and Show Conflict with Sibling Conflicting and suspected Markers based on Mendelian inheritance are highlighted Show Genotype Toggle between Displaying and Not Displaying the Genotypes of the selected node Family 2 2 2 Famip2 Jones Family Ek 1 cu a 4 5 le E Select a family from the currently uploaded pedigree file to view and edit 12 TPOX vee Select a Marker or Locus to view in the Electropherogram Charts Show Color Allows the user to select all colors to view hide all colors or choose a single dye layer Choose a single dye by single left mouse clicking on the icon Zoom In Use the icon to zoom in on the image or hold down the left mouse button and draw a box from the top left corner to bottom right corner around the area you wish to zoom in Zoom Out Use the icon to zoom out on the image or hold down the left mouse button and draw a box from the bottom right corner to top left corner Set Axis The default setting automatically sets the Y axis according to the maximum peak intensity of the samples Two other options are available auto fit the Y axis using peak intensities of the alleles or the user can select the ranges for the X and Y axis 103 August 2013 Chapter 8 Relationship Testing Kinship Analysis ale s Browse by All ColorsD
187. ne nenne nennen 33 CREATE AN SMP FILE doseszsestvexaraszitu nva da du REOR RE RUE REM ARAS 96 CREATE SE 54 4 52 56 58 59 CREATE NEW PEDIGREE ee eee eene ee ee ee ee eee eene eene nens 97 CREATE NEW SIZE STANDARD eee eee eene eee ee eene nene e enun 43 CUBIC SPLINE METHOD EO n ER E EekE ERE xx Fi i 17 CURRENT OLD nnne nennen senses sese aan 131 CUSTOM PANEL CREATION ssscscscscscscscsccscscscscscscscscscscsceses 56 August 2013 Index CUSTOM SIZE STANDARD 41 5 35 D DATTORMAT six sexi PEIUS 96 DATA FILE LIST iis 11 DATA PROCESSING Jiu duke sa ui o ta uk aud seve RE cana OUR aanecebicate 18 19 DATABASE SEARCH xn 6i n IURE SIR 82 92 104 DECIMAL PRECISION a pea ae po p na UNS o na EE 29 DEDUCING MISSING 112 DELETE BIN m 55 DELETE INDIVIDUAL eese eu ana n un nn auus oko anu os 94 113 DELETE MARKER c sia ds ea cn aa ent ege aant et gen 54 58
188. nel will be copied to the selected directory and will also remain in the Panel List and SoftGenetics GeneMarker HID Panel directory The Panel will be exported as an XML file which can be opened with Internet Explorer Microsoft Excel or Notepad Reload Panel Click Reload to undo editing changes to the Panel The most recently saved Panel will be restored NOTE If the user makes changes to the panel and then answers NO to the The Panel has been changed save changes to file the changes will remain in the Overlay View until the user chooses Reload Panel or GeneMarker HID program is closed Sample List The Sample List contains all the samples uploaded to GeneMarker HID in the current project Samples with a checkmark next to the filename will be displayed in the Overlay View Double click the sample filename OR right click the sample and choose Select De Select to enable disable it in the Overlay View Right click any sample in the list and choose Select All De Select All to display all or no sample traces in the Overlay View Sorting Options Sample Name Sorts the samples in alphanumeric descending order Sample Name sorting is the default option Size Score Sorts the samples by the lane size score as it appears in the Size Calibration Charts See Chapter 4 Fragment Sizing Standards Samples with higher scores will appear at the top of the list Overlay Trace Ihe Overlay Trace displays all selected samples in the Sample List The Ma
189. nflict Toggle between Show Conflict with Parents and Show Conflict with Sibling Conflicting and suspected Markers based on Mendelian inheritance are highlighted Li Family 2 2 2 Famip2 Jones Family Select a family from the currently uploaded pedigree file to view and edit Maker 12 TPOX Marker Select a Marker or Locus to view in the Electropherogram Charts 98 August 2013 Chapter 8 Relationship Testing Show Color Allows the user to select all colors to view hide all colors or choose a single dye Choose a single dye by single left mouse clicking on the icon Zoom In Use the icon to zoom in on the image or hold down the left mouse button and draw a box from the top left corner to bottom right corner around the area you wish to Zoom in D 5 gs y Zoom Out Use the icon to zoom out on the image or hold down the left mouse button and draw a box from the bottom right corner to top left corner E Set Axis I The default setting automatically sets the Y axis according to the maximum peak intensity of the samples Two other options are available auto fit the Y axis using peak intensities of the alleles or the user can select the ranges for the X and Y axis Browse by All Colors Displays a comparative view of sample electropherograms by dye color Individual samples can be selected from the drop down menu Li What to Expect Mendelian inheritance is guided by Mendel s two basic laws Segrega
190. nit is known and GeneMarker HID will place Bins at the specified interval Auto Binning Fixed Bin Width Check this option to enter the number of basepairs on the right and left of the center of the Bins If 0 5 is selected as the Bin Width then the total Bin range will be 1 0 basepairs Auto Label When deselected the Bins will be automatically labeled with the basepair size of the Bin position to the nearest tenth of a basepair If selected the number will be rounded up to a whole number value Update Marker Alleles Marker Name pi Opd M Nucleotide Repeat Auto Detect C Set by Manual 7 Fixed Bin Width 15 M Auto Label Boundary bps 351 Ta 427i Edit Group Allele To associate Bins with a different Marker hold down CTRL key and left Change Marker click and drag across peaks at the edge of a Marker A light blue hashed box will appear Right click in the hashed box and select Change Marker The Edit Group Allele box will appear Select New Marker and a pre defined name will appear Use this Marker label or create a new name and click OK The highlighted Bins are now incorporated into the newly created Marker Delete Marker C Use Existing 25742 New Marker D135742 1 mm Right click the Marker bar and select Delete Marker OR right click the Marker name in the Panel List and select Delete Hot Key DEL Bin Opti
191. nt child Siblings Half siblings Uncle Nephew Cousins and Grandparents P2 2 Pray 91 Poy Po Where Po xy Probability of 2 alleles IBD T given the genotypes of sample x and sample y Pi xy Probability of 1 alleles IBD T given the genotypes of sample x and sample y Probability of 0 alleles IBD O given the genotypes of sample x and sample Kinship Formula Transition Matrices Identity by Descent IBD 4 Po for AA AAs AA each relationship category AA 1 0 1 i AA 0 1 0 Do Parent Child 0 1 0 AA 0 0 1 Siblings 0 25 0 5 0 25 Half siblings 0 0 5 0 5 AA Cousins 0 0 25 0 75 AA n ps 0 A A Uncle nephew 0 0 5 0 5 E aa aaa 0 0 5 0 5 T as 05p a 95 Grandchild AA 0 p p Relationship Testing m kinship Formula a Paw x Do P tx x Pp xv x Po Where gt Identity by descent coefficients for sharing 2 or 0 alleles 1 0 b P xy The probability of genotype v given genotype x with 2 of their alleles IBD P xv The probability of genotype v given genotype x with of their alleles IBD P xy The probability of genotype v given genotype x with 0 of their alleles IBD Formulas for each possible combination of alleles derived from stochastic matrices of Li and Sachs Pa Pg Pc Pp Probability of that allele for a given population AB AB P24 9 5 p pg 2 PoP Pg AA AA o PPa Popa AA
192. nted by a row so that the Marker name may be listed several times according to the number of alleles in the Marker This option is only active when a Panel is applied to the data Columns Click the Columns button to open the Set Peak Table Columns box All column options are listed in the All Columns field on the left The columns currently being displayed in the Report Table are listed in the Selected Columns field on the right Selecting Columns Single left click options in the All Columns field and click the Add button Gammen w to add the column option to the Selected Columns field Hold down CTRL gue n or SHIFT key to select multiple options then click Add Click the Add a All button to move all the options in the All Columns field to the Selected As Columns field De selecting Columns Single left click options in the Selected Columns field and click Remove to move the column option to the All Columns field Hold down CTRL or e SHIFT key to select multiple options then click Remove Click the NI Remove button to move all the options in the Selected Columns field to the All Columns field Click OK in the Set Peak Table Columns box and the Allele Report Settings box when finished The options in the Selected Column field will be displayed along the top of the table in columns Show Only Uncertain Alleles When selected displays ess eas puse pie pe Suan rers caseus only th
193. nvert Mac files to PC files at http www appliedbiosystems com support software 3100 conversion cfm Validation Version The validation or trial version of GeneMarker HID can be installed on as many computers as you wish The trial period expires 35 days after installation of the software Installation 1 Insert the SoftGenetics CD into the CD ROM drive If your computer is not set to automatically open a CD navigate to the optical or CD ROM drive on the computer and open the directory Double click the GeneMarker HID Setup executable file EXE The Installation Wizard will launch Click the Next button in the Welcome window Read the SoftGenetics End User License Agreement and click the I Agree button in the Read Me File window 6 Select Install GeneMarker HID Recommended in the Select Program window and click Next 2 Soll Geeet ix DernrMarker V2 2 0 y xj Welcome to GeneMarther V 7 0 Setup pogan Ths wil ental GeneM ace V 2 11 on your compuler It ib strongly tecommended that you ext Window prog ara runeang the Setup Progen Cick Cancel to qut Setup and close any program you hove nerwe g D Ned lo wih the Setup program Im progam protected by um and bhe PUR PIS VAnNNS T Urnaut dieproducion or of the pogam o ary U wied reprodu ponnn of t may velt in severe cvi and crea penales avi wil be procecuted to the maam extent possible ande law
194. oct cd ues s i xm CIN ceed at DELE cec Luc t seek aces 74 CHAPTER 7 MIXTURE ANALYSIS 0s gestus a d Oves sans does Qual Ses ex aS CO 76 OER VIEW nice tem se DIE E ase cae DAN ED ME CIC neat ID EDU UE 77 PROCEDURE NE a a E 77 Identify tbe Presence ofa MIXEUFE ic E im 77 IVECO PII C T ea upaanee 79 assu bs icu scio hcc T 80 WHAT TO EXPECT MIXTURE ANALYSIS AND DATABASE SEARCH ccsccscsecsecsececcececseesecuccessessecsececceseeseeeceesesseeseeeseusoeeas 81 With Reference Fre 5 aei tex eu pite dau r ced UN Eo cae IE 81 Without dq Rer Trence Fl Ong aad oto noid nies 82 eel erue cM n 86 SAVE AND EXPORT RESULT TABLES i00 oci AE redu TRO Desi o Un dataset ecd vds Do ares c ec 87 IVIIXTURE ANALYSIS EQUATIONS 52 ei ded Un Ee D va oe eb e vade ru abes 87 2 August 2013 Table of Contents CHAPTER 8 RELATIONSHIP TESTING uds eee inia eva Te ER Seek ONES enR S NES eos UNDE SERT e iR ero E ex eEEE E IS ERES aa Tee ee RE REP EE CEE NE 92 Qa RR 9M Q 93 Prote UTE cT 96 Icons ano FUNCOMS bru e ia Ld V E 98 odo dor
195. of nucleotides in each repeat unit of the alleles and place Bins at the appropriate interval Set by Manual Select this option if the number of nucleotides in the allele repeat unit is known and GeneMarker HID will place Bins at the specified interval Auto Binning Fixed Bin Width Check this option to enter the number of basepairs on the right and left of the center of the Bins If 0 5 is selected as the Bin Width then the total Bin range will be 1 0 basepairs Auto Label When deselected the Bins are automatically labeled with the basepair size of the Bin position to the nearest tenth of a basepair If selected the basepair size is rounded up to a whole number value Edit Marker Double click the Marker bar OR right click the Marker bar and select Edit Marker The Edit Marker box appears Adjust parameters and click OK NOTE The Edit Marker box can also be accessed by right clicking the Marker name in the Panel List and selecting Edit Marker Parameters Marker Name Edit the Marker Name field to change how the Edit Marker Marker will be labeled in the Panel Nucleotide Repeats Use the Nucleotide Repeats drop down menu 1 6 or enter a value into the field to set the number of basepairs PUE ns expected between each allele in the Marker Nucleotide Repeats x 4 zi Boundary To move a Marker left or right hold down SHIFT key Boundary 1184 To 1738 and left click and drag the Marker bar To adjust
196. ofile and one single source profile that is a potential contributor to the mixture Null hypothesis this individual is a contributor to the mixture sample Alternate hypothesis some other random unrelated person from the population is the contributor Probability of Alternate hypothesis p for homozygotes 2pip2 for heterozygotes D Examples minem pe eee ue repr Ee t PETRI us D3S1358 Alleles Frequency m de im Zo Wm 16 0 2315 foes ns m jme 17 0 2118 Em rums e ie LR 1 2 x 0 2315 x 0 2118 10 197 Hee eee Bem D18551 Allele Frequency HO we u d aa 12 0 1276 Euer won meas S Lu LR 1 0 1276 x 0 1276 61 42 When two potential contributor profiles are available the following scenarios may be tested Scenario 2 Null Hypothesis Mixture is from A and B Alternate hypothesis mixture is from person B and one unknown person unrelated to A The project contains a mixture and two single source potential contributors Both single source samples are selected and person A is contested In the column for Major contributor allele combinations we see there is only one viable allele combination for the major contributor person A 12 14 or a b Example for D851179 Alleles Major contributor a b 12 14 Minor contributor a c 10 12 Mixture 10 12 14 PowerPlex 16 US Caucasian allele frequencies 10 0 1020 12 0 1454 89 August 2013 Chapter 7
197. ogram The Configure Registration window appears Click Register Now to register the local license Click Register Local Text based Key from the Choose Registration Method dialog box Proceed through the Registration steps as described in the Registration section above Launch GeneMarker HID and begin analysis P up ot OY N Network licensing Option The network licensing version of GeneMarker HID can be installed on any computer in a network configuration SoftGenetics uses the License Server Manager LSM to control the number of concurrent users accessing the network licensing option of GeneMarker HID v2 00 and above LSM uses text based registration no hardware is required Both software components are installed from the same EXE The computer where License Server Manager program is installed is considered the Server computer Computers on the network other than the Server are called Client computers 7 August 2013 Chapter 1 Installing GeneMarker HID Installing License Server Manager will require restarting the system to complete installation Please save all work and close all applications before installing LSM 33 SeltGenetics CemePiarker V2 2 0 Install License Server Manager piii o 1 Insert the SoftGenetics CD into the optical or CD ROM drive If your S RUN E T computer is not set to automatically open a CD navigate to the optical tl Leni ver Mni or CD ROM drive on the computer and open the directory T eii
198. on Select to display Size Height and or Area of the peak all within the same cell Parentheses separate the peak statistics from the Allele Name Only enabled when Vertical Orientation is selected I Show Rejected Low Score Alleles Hide Extra Sample Names Bees 238 22 5 28 327 ae b pszaos scr B Josz moase k Josz mosser E foen je Josz cosse ep pessos scr e rosser 18 Orientation Horizontal Sample names appear on the left in rows and Markers appear at the top in columns Vertical Sample names appear in the far left column in rows with Markers listed in the second column Alleles in order of basepair size appear at the top in columns 23 D05 pesceoe ser s fesz rosser ee Show when no allele call Allows user to specify a symbol or short word to enter in the cell of the allele report when there is no peak no amplicon at that marker If not selected these cells will be empty in the allele report amp aRRORRERRIRERREREREREERRERRRRRREREER 67 August 2013 Chapter 6 Reports and Printing Show Only Uncertain Alleles When selected displays only the peaks with Quality ranks of Check yellow and Undetermined red Show Rejected Low Score Alleles When selected the peaks with peak scores be
199. ons Create Bin To create a Bin position right click in the electropherogram at the exact position to place the new Bin Select Insert Allele The Allele Editor box will appear Adjust parameters and click OK Allele Enter a name for the Bin All peaks that appear within the Bin will display this value in the Allele Label in the Main Analysis window Size Indicates the basepair position of the center of the Bin Boundary Indicates the range of the Bin on either side Left and Right f ele Editor of the Bin center Marker Select which Marker to associate the Bin with The Markers to Allele 116 ok the right and left of the Bin position will be displayed as well as the Size 1163 Cancel option to create a new Marker for the Bin All Bins must be associated Boundary Left 05 15 with a Marker Marker f BAT26 C Penta D 54 August 2013 Comments he Control Gene SS NS Chapter 5 Panel Editor Comments Free form text field to associate a comment with the Bin Control Gene Select Control Gene if the Bin contains a major peak in the Ladder samples Bins marked as Control Gene will display the Allele Label below the Electropherogram trace of the Ladders in the Main Analysis window Bins marked 0 will not display an Allele Label even if a peak is present Edit Bin Right click the vertical grey bar in the center of the Bin in the Overlay Trace Select Edit Allele and the
200. ons Settings box and will be applied to the data without having to re analyze the data with Run Wizard or Auto Run Apply to All Apply to Current Re analyze Individual Samples To re analyze an individual sample dye color or marker click the Call Allele icon in the main toolbar The arrow next to the icon opens the drop down menu with additional options Click an option from the drop down and the Recall Allele box will appear Adjust parameters as necessary and click OK The new parameters will be applied All Samples Freel Allri Call the Mater Applies the new parameter settings to all samples in the dataset E similar to Run Wizard and Auto Run l Arkkhnal 5 pusta DE mim m Open Samples Alas Call Applies the new parameter settings only to samples that are PARES filie Cn Tarn checked in the Sample File Tree AED 6 i Peak Datechon Tiazholi Asie E vali nhon Hin inier hn z br br hi mm x Peak FESE Current Sample i M Percerdagem 2 Biol Hx Applies the new parameter settings only to the sample highlighted in the Sample File Tree Shem eos Call the Dye Applies the new parameter settings to the dye selected in the ESE Recall Allele Call Allele by Dye field Call the Marker Applies the new parameter settings to the marker selected in the Recall Allele Call Allele by Marker field 19 August 2013 Chapter 3 Main An
201. ontributors to the mixture Screen saves of samples and examples of comments are presented below Please go to the Mixture Analysis Likelihood Ratio and Hypothesis testing section for example calculations 84 August 2013 Chapter 7 Mixture Analysis BB a m e amen os E etmene Meow Teo Conhibulon DEIA nm aa naw fi nsa VLA amp META SS in meri Cri A a urn Hz iem ie ny ES uy LUND Li 222 xim a hee du am A PM E AREE TAE 1517 pneu id n T ues ue T 15 12 13 OSB aii Pa il aoe i 0399 um oe mhie 15 TON Dew MN DRE FRsi30 Oa T d Y hi od 88121 ig aka 1713 DEG DER i214 a joa ime vn EE VEO o rH A DM ki y Erie an mni de AK OSE mex i cT LN 8 1 bei ai TE t ENS Qr qan ri ALSE uir 18 265 Dr1954 71 1413 LEAT nas Y a waT ie De KI uu WAV Mamet qme E i f aa KY EH mes A A Am ni E 47458 pies ALI 1215 nM oma m nas a E Ama ma TW EENEN wy My om AMEL n5 og i pame a0 o5 um D Fk mu 22 ux F s A 1 1 Ud naa MEA ma aa m UE 100 aM
202. oolbar Choose a directory enter a filename ProjectName AlleleReport is the default and save as an Excel xls or tab delimited Text txt file To export only selected cells in the report table first select the cells by left mouse drag across the cell range or hold SHIFT key and select cells Right click on the highlighted cells and select Copy Hot Key CTRL C The information is saved to the Windows clipboard and can be pasted into any common word processor or spreadsheet program like Microsoft Excel The row and column headers for those cells will be copied with the highlighted cell information Menu Options The following menu options can be found in the menu bar of the Main Analysis window File Menu The File menu contains functions for opening and saving raw and processed data Ga Open Data Open Project Open Data Reopen Project Launches the Open Data Files window where the user can select raw data files for upload into GeneMarker HID Accepted file formats include fsa ab1 abi scf rsd Save Project esd smd smr See Chapter 2 General Procedure Close All Open Project Exit Opens a folder search window where the user can select to open previously saved SoftGenetics GeneMarker HID project files sgf sfp Re Open Project Saves the last four projects that were opened by GeneMarker HID and allows the user to launch any one of those four projects directly Save Project Saves a SoftGenetics GeneMarker H
203. or a profile deduced from the mixture sample matches profiles and calculates the LR Tools Menu The Tools menu contains the Panel and Size Editors in addition to other helpful Took Help modules Chapter 4 contains details on the Panel Editor and Chapter 5 contians panel editor details on the Size Template Editor Please see Chapter 9 for details of all additional U Size Template Editor tools Positive Control Template Editor File Conversion Panel Editor Project Comparison Provides a variety of tools to adjust edit and create control Panels See Chapter 5 Pedigree File Name Match s File Name Group Tool Panel Editor Convert Text to Binary Files Replicate Comparison Size Template Editor Allows the comparison of sample files against a selected size standard to modify and save the size standard for future use or create a customized size standard See Chapter 4 Fragment Sizing Standards Output Trace Data Export Electropherogram 3l Magic Wizard Show Last Event Positive Control Template Editor This menu enables the user to enter positive control genotypes making them available in the third screen of the Run Wizard for automated positive control concordance See Chapters 2 General Procedure and 9 Additional Tools File Conversion This tool allows import of time and distance files from custom genetic analyzers for use with files formatted by the Convert Text to Binary File tool Project Comparison Allows
204. or linearity of lane analysis NS Show Chart Table loggles display to show only the Peak Table the Peak Table and Electropherogram or just the Electropherogram Save Peak Table Exports the Peak Table as an Excel xls file or tab delimited Text txt file gd Call Allele Call alleles by sample s by marker or by dyes Permits slight modifications to the samples without having to activate Run Wizard again Settings to change include Peak Detection Threshold Stutter Peak Filter and Peak Score Threshold Marker Drop down Menu Marker ellw 1 Allows the selection of a marker to view This is available after the samples have been compared to a Panel Event Log Displays each lane s processing success or failure ati Magic Wizard Activates the Start Your Project Run and or Report dialog boxes Report Table Icons The icons are located directly above the Report Table Report Settings Allows the user to customize Report Table display settings Save Report Exports the Report Table as an Excel xls file or tab delimited Text txt file Customize Bin Column Bin 35 August 2013 Chapter 3 Main Analysis Overview Allows the user to select which bins to include exclude in the Report Table Additional Analysis Options In addition to the Main Analysis window there are two other display options in which the sample data can be viewed Browse By All Colors and the Profile Comparison View Browse By
205. ositions When Size Calibration Charts is closed the Size Match Score indicators next to the filenames in the Sample File Tree in the Main Analysis window will be updated Copy Current Calibration Data When selected the frame position and basepair position of the green triangle peak indicators for the selected sample will be copied to the Windows clipboard and can be pasted into a spreadsheet or word processing program such as Microsoft Excel or Word Calibration Plots The Calibration Plots chart the migration linearity of the ILS fragment peaks for each sample The charts plot the peak basepair positions on the y axis as a function of time raw data frame numbers on the x axis As the linearity of the line decreases so does the Match Score for the sample Incorrectly identified peaks will result in a low Match Score Double click a Calibration Plot to select the sample in the Sample List and display the sample in the Sample ILS The currently selected sample filename will appear red in the upper left corner of the Calibration Plot Procedure After a Size Standard has been chosen and the data is processed by the Run Wizard the Size Calibration Charts can be used to correct improperly sized samples 1 Click the Size Calibration Charts icon in the main toolbar 2 The Calibration Charts window appears 46 August 2013 Chapter 4 Fragment Sizing Standards eS Select a sample to edit in the Sample List The sample s
206. otype is unknown The node is displayed as an empty square or circle Unaffected The individual does not show signs of the expected phenotype The node is filled in white Affected The individual expresses the phenotype The node is filled in with diagonal hashed lines Sample File Select the individual s sample file from the drop down list Only samples in the current dataset will be available If no sample file is chosen the node will be grayed out Additionally drag and drop a sample from the Sample List onto a node to associate the sample with the individual node Add Family Members To add family members to the Pedigree Tree right click a node and select Add Mate or Add Child The Add Mate or Add Child box will appear Enter the individual s information and click OK Display Conflicts Once a family is created in the Pedigree Tree GeneMarker HID will automatically identify any Mendelian inheritance conflicts between Parents and Children or between Siblings Click the Show Conflict with Parents Siblings icon in the toolbar to alternate between the two modes Nodes with conflicts will appear red Mouse over the red node and a list of Markers or loci with conflicts will appear Single left click the Marker name and the family members electropherograms will appear in the Charts tab on the right The conflicting Marker will appear red 44 Det Additionally suspect Markers are indicated in the Pedigree Tree by a question mar
207. p down menu hs 7 5 nes xa amm 1x3 D165533 i 13 RIMIS tiewe sew i ib 00000000 4 Select Allele Frequency of the ced E js mM rcc appropriate population from the Tools lotacan lia 2 206954 Fielahanihigi Corning drop down menu oe qb ond Hi Uic d ss 5 Diod ia Fs a pa P 200000 Poeni hr 5 Select Kinship Analysis Tool use drop bes i n z Loos E MCN down menus to select individuals for piss ji i5 5 n 43570 D Hadas Uz1511 a Fe al 3A 1 15225 a UT comparison UM Ai lees cle Hegh J F mine Comm 6 Select the desired relationship level and Proc Scar 2 MEO Dirandearent Crandchild likelihood ratio probability or both at the kinship analysis settings Probabilities for the occurrence of the genotypes within the population having a specific relationship or being unrelated for each locus and all loci combined are displayed in table form 7 Likelihood ratios for each locus and combined likelihood ratio of a related parent child sibling half sibling are presented in table form Icons and Functions JA Relationship Testing Relationship Testing Main Drop down menus Fade DetaBsce Tools Select from File DataBase or Tool options Relationship Testing Tools include M tenga Family Group Tool for automated pedigree trio drawing Allele Frequency for major US populations Budwole 2001 et al lY m a Family Group Tool M Allele Frequ
208. play the sample filename in red font in the Sample File Tree Default is NC 30 August 2013 Chapter 3 Main Analysis Overview NOTE To implement a change in the Identifier fields right click any sample in the Sample File Tree and select Set Sample Type Auto Identify Report Settings The Report Settings tab allows users to select how data is displayed in utn the Report Table Automatically Re Sort Report Check this option if you would like GeneMarker HID to automatically re sort the report every time you modify alleles Un check this feature if you want the report to remain sorted until you choose to re sort Automatically Scroll Charts to Alleles When Selected in Report You may choose whether to scroll to alleles in the trace when selecting pas iet Y a eae cd the allele in the report Leave this feature on to have the software automatically call up alleles in the trace when you double click on them in the report Show Disabled Samples in Report GeneMarker HID identifies samples that failed during electrophoresis or size calling The default setting excludes the disabled samples from the report The Bee Caes option may be selected to have failed or user disabled samples to i be identified in the report Add Prefix to Saved File Name of Ladders and Controls Automatically adds the prefix Ladders to the allelic ladder file names when saving report and Controls prefix to any samples designated as PC or NC s
209. plays user defined peak statistics Sample names are displayed in the far left column in rows and the Marker names are in the column adjacent to the sample names In columns at the top of the table are the selected peak statistic information labels The column options available in the Peak Table Report Style are similar to the options available in the Peak Table that appears below the Electropherograms See Chapter 3 Main Analysis Overview for column option definitions Features Options Size Range bps When selected allows the user to define a specific 69 August 2013 Allele Report Settings Report Style C Allele List Forensics C Bin Table C Allele Count C Sample Name File Name r Orientation Options Size Range bps v Abide By Panel Grouped by Markers Columns Ak Show when no allele call Show Only Uncertain Alleles Vv Show Rejected Low Score Alleles Hide Extra Sample Names Cancel Chapter 6 Reports and Printing basepair range Only the peaks within the range will be displayed within the Report Table Abide By Panel When selected the table will show only called alleles within Panel Marker ranges This option is only active when a Panel is applied to the data Grouped by Markers When selected alleles within the Marker will be listed one after the other in the columns at the top of the table When de selected each allele will be represe
210. ple Name and Score columns will appear in the Sample List The basepair size position of the disabled peak is reported for each sample If the disabled peak is at the beginning or end of the Size Standard no basepair size position will reported 44 August 2013 Chapter 4 Fragment Sizing Standards Size Calibration Charts Sample List Calibration Plots Sample ILS Disabled Size Statistics Size Standard Trace LIII 2 euo ie nm Fite I 04 woo 4 s ittm LADO bee M irh t E e m ite Compton ommo B Disabled Size Statistics If Sizes were disabled in the Size Standard see previous section Size Template Editor then the Disabled Size Statistics table will appear in the bottom left corner of the Size Calibration Charts window The average basepair position the standard deviation and the difference between the maximum and minimum basepair positions across all samples are calculated for each ILS peak matched to the disabled peak s position No statistics will be calculated for disabled peaks at the beginning or end of the Size Standard Size Standard Trace The Size Standard Trace displays a synthetic trace of the selected Size Standard Enabled Sizes are red disabled Sizes are grey Each peak in the Size Standard Trace represents the expected basepair size of peaks in the sample ILS Sample ILS The Sample ILS displays the currently selected sample s ILS
211. pter 3 Main Analysis Overview The red horizontal line seen here in the figure on the right is to alert analysts to trends in the data These areas of the data have a more elevated baseline or noise to signal ratio often associated with poorly resolved peaks than the nearby regions of the trace which sometimes masks very minor peaks Peak Table The Peak Table can be displayed below the Electropherogram by clicking the Show Chart Table icon in the main toolbar Right click in the Peak Table and select Show Columns The Show Columns fly out appears with column options Dye Indicates the dye color of the peak Gf ladder ATI bd x I I Ji i i 131 UU Uu WLU we POEL Sy Lal AU WA 7 1 is 1009 23 is Mill 8 Oye irren 7 Ud Sees 2 t 14 2 t Lo Lit t 24 11 15 t Size Indicates the basepair size of the peak x axis Height Indicates the peak height in RFUs y axis Height Ratio The value obtained when the peak s height is divided by the height of the highest peak in the dye color or Marker Va Height lt a 4 Fato Area Indicates the area under the curve of the peak The area Ares 0312 Edit Allele Ar Rabo m 5 x M e ee calculation begins and ends along the x axis as indicated by wi 5 pao M on rcm fin 1255 178 px Dui the Start and End columns of the Pea
212. pter 9 Additional Tools Automated Control Concordance Filename Group Editor Output Trace Data Project Comparison Convert TXT to Binary Export Electropherogram 116 August 2013 Chapter 9 Additional Tools Automated Control Concordance Positive Control Template Editor 1 4 5 Tools Positive Control Template Editor to launch dialog box Import Genotypes dropdown menu Select from functions to Add new positive control samples Edit or Delete files from samples using Positive Control Template Editor w X Cancel D195433 14 15 vwA 18 TPOX 8 8 D16551 15 13 AMEL X A D55818 11 11 o 23 24 r Positive Control Template Editor Genotypes 1 D851179 2 D21511 3 D75820 4 CSFIPO 5 D351358 THO D138317 8 D185533 3 D291338 10 D135433 11 12 TPOX 13 D18551 14 AMEL 15 D55818 13 30 10 10 14 11 11 13 14 17 15 11 Positive Control Standards PC NISTADENTIFILER fsa Allele 1 Allele 2 13 30 11 12 15 3 3 11 12 23 15 18 13 11 Import Genotypes from Sample E Delete Select the appropriate positive control file from the dropdown menu in the run wizard Summary Additional Settings Hdentibiler Analysis Det sod aphoni ha ha Gen arce type Alcides PU hd uin imd f ifm Posten Contd PEHIST IOEHTIBREER f
213. r Use this function if the genotype is only available from an archived file the fsa or hid data file is not available Format the txt tab delimited file as in the example below the exact format is required for the program to recognize the genotype The file must be saved as txt tab delimited AA3 f 18 4 A B D E F G H I J K L M N a 1 AID D8S1179 D8S1179 D21S11 D21S11 D7S820 D7S820 CSF1PO CSF1PO 351358 D3S1358 0135317 D13S317 0165535 0165553 D2J 2 Sample 157 11 13 28 32 2 8 10 7 10 15 15 7 9 3 12 14 10 11 3 Sample 156 12 13 27 28 7 8 8 11 15 14 7 7 12 12 10 11 4 Sample 155 11 11 32 2 32 2 8 8 10 10 15 15 9 3 33 14 14 10 11 1 Open a saved project in the main analysis screen ET Mater zj sen 2 Select Applications Relationship Testing ame 3 Select Tools Allele frequency and select e iE Wm the appropriate table M P wees ae m 4 Select File New Pedigree tt T he 5 Fill in the sample identifier or name and ios click on Manual Input and the open folder Doves j Icon eet 2 NEC 6 Use the open folder icon in the Genotype 777 i Editor to add genotypes from a txt file C Fom Database ire z 7 Select the desired genotype for the search ess right click on the open area at the left to a Remove Selected Files or to Clear all f
214. rameters in the Project Option Settings window This three tab window contains settings identical to the Run Wizard Adjust settings in the Project Options Settings box before selecting Auto Run See Chapter 2 General Procedure NOTE Auto Run does not need to be selected after adjusting the Additional Allele Evaluation Peak Score settings The changes will automatically be applied when the Project Option Settings window is closed Project Comments Allows the user to write free form comments regarding the analysis These comments are saved with the project file and can be displayed in the Print Report Applications Menu The Applications menu contains individual modules for specific data and analysis types These modules present advanced features and reporting options necessary for the particular application Pedigree Applications Tools Help Display and check genotype calls using a pedigree chart Data must be run with 43 Pedigree the correct size standard and Panel prior to using the Pedigree function See Export CODIS Chapter 7 Relationship Testing Profile Comparison View Export CODIS Relationship Testing Developed for forensic scientists analyzing short tandem repeat fragment data Exports the CMF 3 2 xml and CMF 1 0 dat files for upload into the FBI s Mixture Analysis CODIS database See Chapter 6 Reports and Printing Profile Comparison View Allows the user to graphically display any combination of samples and dye
215. re on the far right of the trace Haw Data Analysis Window Sample Tree Synthetic Gel Image Raw Trace Data GeneMarker HID Untitled File View Project Applicatio Lint TRUE Hed amp Rew Data Mb5cace B MixX05casel ongeene B MOWScere1 B MiX Sc 5 BM BM B ngeen IB M BM E M DSN BM 1 B LO PowerPlex1E LADDER isa B NC Poweftex 1E nc Isa 5 PowePlex Ica LUI Hil ase 0 5 00 200 100 00 900 00 00 600 500 00 90 200 100 t Projec gt Page Up Page Down Sut n lary New 15 sample PC error 0 0 NC error 0 0 Ladder error 0 0 led 0 SFlagged 0 Main Toolbar Icons s gt x s gt Spike Removal Removes peaks from voltage spikes caused by micro air bubbles or debris in the laser path This option is selected by default in the Run Wizard Saturation Correction A synthetic peak is created based on peak shape before and after saturation The results of these will be less accurate than that of non saturated peaks This option is selected by default in the Run Wizard Smooth This function smoothes the baseline by eliminating smaller noise peaks This option is selected by default in the Run Wizard Baseline Subtraction Selecting this option will remove the baseline completely so that the Y axis will be raised above the noise level This option is selected by default in the Run Wizard Auto Pull
216. reopen the files in Pedigree tool follow steps 1 10 of the Upload Previously Created Pedigree section above Save Pedigree Tree I qe i Extended Pedigree Files All personal information is saved and retrieved with the pedigree smp and dat files including affected status and deceased Extended family trees can be built saved and added to as additional information becomes available Right click anywhere in the Pedigree Tree Select Export Bitmap The Save As window appears Enter a filename choose a directory and click Save The Pedigree Tree will be saved as an image file BMP Save Pedigree File Pedigree File Pre Ped Ic Users SoftGenetics Desktop T estPed pre iw Individual S ample Accordance File SMP D XLIserssSoftGieneticssDesktopXT estPed smp iw Loci Description File dat C Users SoftGenetics Desktop T estPed dat Auto Load Ok Cancel 97 August 2013 Chapter 8 Relationship Testing M Pedigree Edit CA ser SoftGenetles Desitop Relationship Testing New RT peojects testing phenotype saves pat Sgen nodelContested dine descent afe AE mici J ea T iz f Fart 1 Fen Foret oo Mather Al Markers Spach j A lt Samples Chats z No Same ES 3 Bi PAT 1 nh 2 PAI Fisa 7 1 PAT 1 M fre S SS lay 3 B PAT 2 Clean WY 9 13 10 cl m SN Si p Li Bs 5 FAI 2
217. rker bars appear above the electropherogram and the Bins appear within the electropherogram as brackets at the top and bottom The center of the Bin is indicated by the vertical grey bar in the electropherogram only in Panel Editor The Overlay Trace view can be changed by clicking the Trace Mode icon in the toolbar Other options include Max amp Average and Gel Image Navigation in the Overlay Trace frame is similar to the navigation options in the Main Analysis window See Chapter 3 Main Analysis Overview Marker Options Create Marker Hold down CTRL key and left click and drag across peaks in the Overlay View A light blue hashed box will appear Right click in the hashed box and select Create Marker The Create Marker box appears Adjust parameters and click OK Marker Name Edit the Marker Name field to change how the Marker will be zma labeled in the Panel Boundary The basepair range of the Marker is defined by the range of the light blue hashed box and is therefore inactive in the Create Marker box To edit the Boundary see Edit Marker below Marker Name Green NEW Boundary bps 218 5 Ta 2325 Nucleotide Repeat C Auto Detect Setby Manual 2 PI Nucleotide Repeat 7 M Fixed Bin Width 0 5 Auto Label 52 OK Cancel August 2013 Chapter 5 Panel Editor Auto Detect Based on the peaks present in the Overlay View GeneMarker HID will attempt to detect the number
218. rogram operability It is recommended to accept the default 9 Click Next in the Start Installation window to install GeneMarker HID 10 Click Finish in the Installation Complete window 11 The Installation Wizard will close 12 Bject the SoftGenetics CD 13 Launch GeneMarker HID by double clicking the GeneMarker HID desktop icon OR open the Start menu and navigate to SoftGenetics gt GeneMarker HID the version that was just installed GeneMarker HID program 14 The Configure Registration window will appear Click Register Now to register the local license 15 Click Register Local Text based Key from the Choose Registration Method dialog box Registration 1 The Register Local Text based Key window appears 2 Ifthecomputer GeneMarker HID is being installed on has an internet connection select Online Registration If the computer does not have an internet connection or is connected to a proxy server select Offline Registration Online Registration A B C Locate the Account and Password on the SoftGenetics CD Enter your Account Password and e mail address information in the appropriate fields The Request Code information is automatically generated by GeneMarker HID 6 August 2013 z Com ix s DernrMarker V2 2 SF EB Welcome to V2 7 0 Setup This pogon ental GeneM aer V2 2 ors your compuler It ib strongly vecommended that you eut af Windows pogana belore runeang the
219. rt Styles Sort by Column 27 August 2013 Chapter 3 Main Analysis Overview Select Sort by Column from the right click menu and choose from the fly out menu to sort Ascending or Descending If Ascending is chosen then lesser values will be sorted to the top of the table and greater values to the bottom the table and vice versa if Descending is chosen This option is available with all Report Styles Editing Peaks To edit peaks first left single or double click the cell in the Report Table then right click to see menu options or use Hot Keys Delete Peaks Right click the peak cell in the Report Table and select Delete Peaks Hot Key DEL The deleted peak will be removed from the Report Table Confirm Peaks If a peak is given a low quality score it will receive a Check yellow or Undetermined red Quality ranking To give the peak a Pass green Quality ranking right click the peak cell and select Confirm Peaks Hot Key CTRL M The peak will be marked Pass green Peak Information Hold down CTRL key and click the peak cell of interest The Allele Peak Info box will appear containing information such as Sample Dye Size Marker Allele Score and Comments The information in these fields cannot be edited This option is only available with Allele List Marker Table and Peak Table Report Styles Save Report Table To save all information currently displayed in the Report Table click the Save Report icon in the Report Table t
220. s Right click on the genotype combination table or the likelihood ratio report to copy and paste the results into an existing document Select Export to save tables as a txt tab delimited file Comments that are typed in by the analyst are saved with the likelihood ratio report amp 9 Conii 1 MUADEcase2_yschm Mina US Caucasun alala hegaency welim reference He i only selected com bear LA foe victim as canhar to mele Lamihuler2 Hone Average Majai Mx o SEU Cms TSE punc Mixture Analysis Equations Major Mx A B A B C D User selects whether to use peak height or peak area in the analysis parameters For these examples peak height was selected as an analysis parameter Example using peak height Marker D3s1358 Metre Analyses E Orne i Minos Ma Mee HIM Mine HIM Mx 972 932 972 932 343 526 T ina Female FO 0 69 TCI E ong et Mac F FS 3 351 3122 A Tate o ine Contains 1 1232 1314 iega Mana 0 720 5 a 716 The major contributor accounts for 69 of the DNA in the mixture sample Residual va observed pa expected Observed proportions are calculated from the data Expected proportions are calculated using the methods of Gill et al Forensic Height Ht Ratio Area Ar Ratio Marker Allele Quality Quality Rea 343 035 2840 0 37 D351358 14 Check IMB Sci Intern 91 41 53 table at right 526
221. s will be used for pattern recognition in the sample ILS but will not be used to size fragments in the other dye colors Disable a Size if its position is variable from sample to sample NOTE If the Enabled value is changed in the Size Table you must click another cell in the Size Table before saving the Size Standard or the change will not take effect Enabled Insert Size Right click at the position in the Expected Size Standard frame or in the Sample ILS where the Size should be placed The Edit Size box will appear GeneMarker HID will automatically interpolate the value in the Size field if there are two or more Sizes present in the trace Adjust as necessary and click OK A green triangle will appear at the cursor position indicating where the new Size was placed NOTE The height of the new Size in the Expected Size Standard trace is dependent on the height of the peak in the corresponding Sample ILS trace Delete Size Select Delete Size to remove the Size completely from the Size Standard Alternatively the Size can be disabled by deselecting Enabled in the Edit Size box or by placing a 0 in the Enabled column of the Expected Size Table NOTE Sizing is often more successful when there are many Sizes in the Size Standard Set Value to Column 40 August 2013 Chapter 4 Fragment Sizing Standards Makes all values in the column equal to the value in the highlighted cell Only available in the Expected Size Tab
222. saved for future use by clicking the Save button If you do not want to use a template select the appropriate size standard standard color and type of analysis Use last template will automatically be selected Panel GeneMarker HID comes preloaded with many common kit Panels including Promega s PowerPlex kits and ABI s Identifiler kit Additional Panels can be imported by selecting the Open Files icon next to the Panel field A custom Panel can be created in the Panel Editor tool See Chapter 5 Panel Editor Panel Editor A Panel can be selected from any available from the drop down menu or can be viewed and selected by clicking the Panel Editor icon 15 August 2013 Chapter 2 General Procedure Import a Panel If a Panel cannot be found in the Panel Editor tool it can be imported by clicking on the Import a Panel icon Size Standard GeneMarker HID comes preloaded with many common size standards including GeneScan 500 and LIZ600 A custom Size Standard can be created by selecting the Size Template Editor icon next to the Size Standard field See Chapter 4 Fragment Sizing Standards Size Template Editor This allows the user to check sample files against a selected size standard modify and save the size standard for future use or create a new size standard Standard Color Select the dye color which contains the internal lane standard Analysis Type The Analysis Type option is inactivated in GeneMarker HID Bus Wend
223. shold parameters are only applied to peaks outside of the Panel Markers To adjust settings for peaks within Panel Marker ranges see Chapter 5 Panel Editor Min Intensity Minimum RFU threshold of peak height used for peak detection Peaks below this value will not be called Max Intensity Maximum RFU threshold of peak height Peaks above this value will be flagged with a yellow Allele Label given a Quality Rank of Check and marked with HI Quality Reasoning Percentage Global Max Relative minimum intensity of allele peaks to the 5th highest peak in the dye color used for peak detection Peaks below this value will not be called Load Default Recalls any settings previously saved by the user If there are no user saved settings the program loads the default settings for that particular analysis type Save Default Saves any settings defined by the user that is different from the default These settings can be recalled for consistency of analysis on similar data sets Run Wizard Additional Settings Procedure 7 The Run Wizard Additional Settings box appears 8 Select an Allelic Ladder and adjust the Peak Score Hun Wiad mci Additional MVHCL Analysis Sel ankim oder to Lon diferent oe Abel Ladder Portree Control NIST Poe E Urea sore ag parameters Mele E wae F Aio Pavel Ausiment Peak icma 9 Click OK Biejeci i ITE C pue fu Pass 10 The Data Processing box appears 11
224. show genotype display is used s E Relationship Testing Parameters Launches the Relationship Testing Settings box Options for selected samples or all samples selecting the appropriate allele frequency for the population mutation rate and prior probabilitiy Show Conflict Toggle between Show Conflict with Parents and Show Conflict with Sibling Conflicting and suspected Markers based on Mendelian inheritance are highlighted Show Genotype Toggle between Displaying and Not Displaying the Genotypes of the selected node 8 107 August 2013 Chapter 8 Relationship Testing Family 2 2 2 Famip2 Jones Family Select a family from the currently uploaded pedigree file to view and edit Marker 12 TPOX x Marker Select a Marker or Locus to view in the Electropherogram Charts Show Color Allows the user to select all colors to view hide all colors or choose a single dye layer Choose a single dye by single left mouse clicking on the icon Zoom In Use the icon to zoom in on the image or hold down the left mouse button and draw a box from the top left corner to bottom right corner around the area you wish to zoom in e s Zoom Out Use the icon to zoom out on the image or hold down the left mouse button and draw a box from the bottom right corner to top left corner Set Axis The default setting automatically sets the Y axis according to the maximum peak intensity of the samples Two other options are av
225. single dye color to view in the Sample ILS frame Cycle through the colors by left clicking the icon or use the drop down menu Size Match Automatically places the green size marker triangles atop the peaks of the sample trace and matches it with the selected Size Standard 6 What to Expect Once the Size Standard is created it can be applied to the dataset Save the edited Size Standard in Size Template Editor then exit Size Template Editor If the Size Template Editor was accessed via the Run Wizard Template Selection box icon then the selected Size Standard will appear in the Size Standard field If the Size Template Editor was accessed via the Tools menu then click the Run Process icon in the Main Analysis toolbar The Run Wizard will appear Select the Size Standard from the Size Standard drop down menu in the Run Wizard Template Selection box Proceed through the other Run Wizard boxes and click OK when the Data Process window is complete The Size Standard will be applied 5 gel The success of size calling for each sample is indicated by the green yellow and red sheet next to the sample filename in the Sample File Tree of the Main Analysis window The lane sizing quality is determined by the Match Score which in turn is a calculation of how closely the sample s ILS peaks match to the selected Size Standard If a sample receives a low Match Score the sample will be marked with a yellow sheet If the size calling failed the 43 Aug
226. sociate with the Pedigree Tree node 4 Click OK 5 The individual s node will appear in the Pedigree Tree and the individual ID information will appear next to the sample name in the Sample List 6 Continue to add family members by right clicking at the node and selecting Add Mate Child 7 Mouse over red highlighted nodes in the Pedigree Tree to view Conflicts and Suspect Markers see section above Pedigree Tree 8 Edit alleles in the Electropherogram Charts as required see section above Electropherogram Charts Save Pedigree File New Family New Individual Family Name Smith Mew Individual Person Info Name Woe Gender Male C Female Unkown Affect Status 9 Unknown C Unaffected C Affected Sample File 1273878 30 08 fsa v ck Cancel After the Pedigree has been created and modified there are two options for saving the information save the Pedigree File and or export the Pedigree Tree as an image Save Pedigree File pn 10 In Pedigree tool click the Save Pedigree File icon The Save Pedigree File box appears Click the Save icon next to the Pedigree File field The Windows Explorer directory window appears Enter a filename and select a directory Click Save The directory path and filename will appear in all three fields of the Save Pedigree File box Click OK Three files will be saved to the specified directory PRE SMP and DAT To
227. sociated with several commercially distributed human identiy Kits are included Examples of some of the pre defined Panels include Promega s Powerplex kits and ABI s Identifiler and Yfiler kits GeneMarker HID also offers the opportunity to create a new custom Panel if the pre defined Panels do not include a kit that the user is working with Below is a discussion of how to use the pre defined Panels or create a new Panel with GeneMarker HID s Panel Editor tool The Panels displayed by default include PowerPlex 16 PowerPlex 55 PowerPlex Y PowerPlex Y23PowerPlex ES PowerPlex ESX16 and 17 PowerPlex EXI16 and 17 PowerPlex 18 PowerPlex 21 PowerPlex Fusion DIPplex PowerPlex EURO Pre Defined Panels GenePrint FFFL GenePrint CTTv Globalfiler amp AmpF STR COfiler AmpFtSTR Identifiler AmpFfSTR MiniFiler AmpFtSTR Profiler Plus AmpFtSTR Profiler AmpFSTR SEfilerTM AmpFfSTR SGM Plus AmpF STR Yfiler The pre defined Panels appear in the Panel List Single click any Panel in the Panel List to select it In addition to the Panels displayed by default the user has the option to import standard Panels and Bins Text files Follow the steps below to import Panels and Bins Text files Panels and Bins Files 1 In Panel Editor select File Import ABI Panels from the menu bar pres eS 2 The Import Panels from GeneMapper box appears 3 Click the access button next to the Panel
228. splays all traces of the selected samples in the Samples List one dye color at a time Single click any trace in the Trace Overlay frame and the trace will become bold and the associated sample will be highlighted in the Sample List Max amp Average displays two traces in the electropherogram The darker color line corresponds to the maximum peak height at that position and the lighter color line corresponds to the average of all selected sample traces at that position Gel Image displays selected samples as a synthetic gel image Bin ranges in the Gel Image mode appear as white vertical lines and can be manipulated by holding down SHIFT and dragging the white lines left or right Check Range in Edit When activated the software will warn the user if they set the left or right range of an allele to overlap with another allele This feature will prevent the user from setting allele boundaries too close to neighboring alleles This option is selected by default Major Adjustment of Panel Uses previously defined size and height information located in the Panel file when the panel was previously saved with signal information to identify Marker and Bin positions To be used when a Panel must be adjusted by 1 5 basepairs in order to align with the dataset peaks For use only with panels that do not contain variant alleles 0 in the control column Minor Adjustment of Panel Aligns the center of the Bin to the center of the nearest peak within one bas
229. ssible genotype combinations are displayed in the results table Red font indicates that the combination did not meet the analysis parameters Black font indicates that the 79 August 2013 Chapter 7 Mixture Analysis genotype combination meets the analysis parameters If this option is de selected only the combinations that met the analysis parameters are displayed in the result table Report Save Parameters when Saving Report creates an ini file with the same name as the report providing verification of mixture analysis parameters used to generate each report Mixture Analysis Results Mixture Calculations Results File Name Tree Trace or Report Display Option EY P9 g i Mahou afetakes Make Mas Mrs Maes Mia Aesi Macc HIM Vire HIM Two Corftnbutars 1 1 0951179 111 11 14 ae uo 099 593 1 2 nasn 8 0132 1414 033 065 08 Towee ox exe Coni 1 1 pas1179 1113 1114 093 10a n76 23832 10 31 083 2 aeg 7 815 097 mop 15 on Conbutor 1 Coeur 2 LA 6 30579 59421 11 34 5423 02519 083481 30 31 ans e 64501 035233 81 12737 553470 12 13 25512 1536 042337 057653 8310 076630 83 awe 512 025626 121314 43512 5U499 151673 268 gz73i 851011 85 12153718 123954 76146 uas VARLI 034 1L ons 033 0082 055 n 0032 nomo 09 aaa 32 032 0 0064 x
230. st EEEE 51 PANEL ABLE 2 veia suse siue Sax ed exe sax e bas ss 55 PANELS AND BINS FILES ccccccscsccccscsccscscsccscsccscscsccscscsccscs 56 PASSWORD 129 PATERNITY INDEX PII oe dtex ada edu a ke e a rai da 115 120 PEAK COMPARISON 5 120 PEAK DETECTION THRESHOLD e eee ee ee ee eee ee neon eene eonun 18 PEAK INFORMATION ccccccccccsccccscscscccccscscsccccscscscscscscscsecs 28 30 PEAK MATCHED BY vo sus su eran Ero ER ERR PRAE FERE REPRE REEF REPE 120 PEAK SATURATION aut aou sub P EEUU 16 PEAK SCORE va tcp ae vu cuu ga vate vua vieta yate guo 18 PEAK TABLE 25 131 70 PEAK TABLE REPORT STYLE eee ee ee eee ee nene eene eene 69 PED FORMAT 225x259 5x 2E UK URRIEAR nedensen CESEREAR RR NEN ERRORS MURS 96 PEDIGREE iiis euavyCUs In uE DAR ERA VENEVI VA CU VEA DEUM 32 93 PEDIGREE ANALYSIS suni ados euh e peak rv sata abr ra rat a va 92 93 PEDIGREE DRAWING 111 PEDIGREE FILE ose kn eno a n tn Po in Yo in Pro ai p n oon o an Pekin nnus 96
231. strator LAE Tuganalon Scttizerets 4 Enter Organization Name an Administrator username and password T Aun User Protenton 5 Click OK 6 You are now logged in as the Administrator OPT TI ERT M 7 Click the Add User button to add additional users 8 Click the Access Rights button to set up user type access Organization SartGeneti permissions Des 9 Be sure to select Run User Protection and click OK to exit 10 Login is required to open GeneMarker HID after the User Cate Manager is activated User Manager The User Manager tab displays user information and contains options for creating and deleting users User Window Ji User Manager Admin logged in Jet E Displays all users by name type and creation date User Manager History Settings i User Name User Type Create Time Organiz ation Admin Administrator 4 3 2007 12 58 32 PM Add User Ent zat Lm e onathan nalyst 16 David Anal 4 13 2007 2 16 17 PM RES Keyin Reviewer 4 13 2007 2 16 27 PM Run User Protection When selected users will be prompted to log on with a user name and password When deselected any person can launch GeneMarker HID without a username and password Access Rights Change User Organization SoftGenetics Add User Launches the Add User box where a new username and password can be input This is also where the user type can be v Fen User Pictection chosen A user can be
232. sy way to compare several sample traces at once Procedure CINE 1 Select several samples from the Sample List by placing a s vs uo ex an check mark in the empty box to the left of the filename Dis susc T n 2 Click OK ers 3 The traces will appear in the window to the right DEAE 4 Slide the 2D Offset bar to the right to de convolute the traces Eus 5 Select the Line List tab to open the list of traces present in the 5 viewer Select any trace to bold the trace line Ciim msc 6 Select a marker from the drop down list to view one marker eer at a time 7 Select Show 3D to see the traces in a three dimensional view Denim 36 August 2013 Chapter 3 Main Analysis Overview Icons and Functions Dye Color Single click to scroll through the dye colors Zoom In Out Single click to zoom in or out on the center of the Trace View window Tools All viewing options are selected by default Unselect these options to change the Trace View settings Save Save the Trace View image as a Bitmap bmp file 37 August 2013 Chapter 4 Fragment Sizing Standards Chapter 4 Fragment Sizing Standards Chapter 4 Fragment Sizing Standards Size Template Editor Size Calibration Charts 38 August 2013 Chapter 4 Fragment Sizing Standards Size Template Editor The Size Template Editor is a tool in GeneMarker HID for creating and modifying Size Standar
233. t samples then click the Open Selected button The electropherograms of the selected samples will open in the Main Analysis window Hot Key CTRL F Sample Information Sample Info Opens the Sample Information box A list of Properties appears and includes information like Sample Name Well ID Lane Number Instrument Name and Chemistry The list of Properties varies depending on the file type Hot Key F2 Edit Comments Opens the Edit Comments box Enter information in the Comments field The last ten comments will be stored and can be subsequently selected for future samples The Sample Comments will appear on the Print Report See Chapter 6 Reports and Printing Hot Key F4 Disable Samples Disable Sample Opens the Input Disable Reason box and marks the sample with a red strike put Disable Reason c3 through A disabled sample cannot be selected for display in the Main Comments Analysis window and will not appear in the Report Table if View Preference TENS Options Show Disabled Samples in Report is deselected Select a Comment Template or enter a new comment in the Comments field and click OK to disable the sample Hot Key CTRL DEL PCR Failed Bad Signal Water Sample Add Samples From the main toolbar select Project Add Samples to Project The Open Data Files box will appear Click the Add button to select additional samples to add to the project and click OK The add
234. t tab delimited files or may be copy pasted into an existing report r Fi x2 Seres Chai 3 Raat f Dasal i a a copy Tabs S60 1 0000000 p ls Expert Cabdstion Decal 1313 M 1043 1516 3 is 0 3m ea au 11 E IUE M 1518 Ui INER 2EM ER 1 132 3 3 3 HoH 12 d 1925 320 os SEG 53044 1 15 8 15 gt 5 3 8 X LOE 2228 7 mo3 LER SIEM an g S180 1008400 2330 fee TE i2 13 it Leo 73 73 73 EIEE 13EM 13 75 lt q j3 item 78 n amp INE 34d 14 13 Tq od 12 1h 1 UE 13 wiz 1219 SEEN BED i it 10430 7119 11 42 1243 inp Ed is id 3 LEO 02 03 GE 22189 A 5 n d 812 12 12x54 T w m TUE MW 1216 1114 234p TIED gt 13 1014 1314 iie Ei 1 007 3 812 1213 1 16 00 1517 1817 1517 2E 504 E 18 44 E 0e 10000000 28 28 x 5 IAS EO 1516 914 gis LZt4e LEN id a 1517 1516 1518 HE 15 18 TEMO 0E 192 1320 2025 TRER 73x40 73 4 1064 2328 2 2021 seis 534 e 5 in LEER 7531 ya E Ime VEN 25 20 MEME 74 73 74 fem 14 13 nuege 7 211 SEEN LET 5 S H 1043 13 1412 1513 IAEI SAEN 15 3 LEE 3 Wiz 1243 IEE BSED Y DEM 1012 1113 114 anew En r4 e E 812 12 LER 25652 1 amp 4 1116 i314 BRED LUE QU 12 an 1314 24 speg Ie E13 812 124 EIS ED EON 1917 Mor Ur ISO TIEN August 2013 115 Chapter 8 Relationship Testing Chapter 9 Additional Tools Cha
235. te Comparison Tool is designed for projects in which multiple replicates of each sample have been uploaded This tool compares replicates from the same sample to each other with the goal of determining a Consensus Genotype The results of this comparison are displayed in the tool s Report Table If a comparison resulted in a conflict e g one replicate had the genotype 11 12 and its counterpart had 11 13 the marker is flagged The user has the ability to address these flags by entering the deduced genotype through a dropdown menu Replicates must be grouped prior to utilizing the tool This can be done in the in the tool itself Edit file groups icon or via the apply sample groupings option in the project menu of the main analysis window Procedure Replicate Comparison Setting 11 Import raw data files 12 Process data using the Run Wizard 13 Use the Main Analysis Window and Report Table A Maker Name Allsle Name je Meer Quality to review flagged size and allele calls and to make Dye M Score any necessary edits 14 Group replicates using the File Name Group Editor Marker Name Peak Size project apply sample grouping 15 Select Tools Replicate Comparison Tool 16 Choose comparison settings or use defaults 17 View results in the Report Table 18 If desired export results Peak Matched By Peak Compare Items r o alee are Deleted Size Confirmed Height M Commer t
236. the basepair UR me m KS w range over which a Marker is located hold down SHIFT key and i o 2 mouse over the edge of the Marker bar until a double headed arrow appears then left click and drag the Marker edge to increase MiHn ete 50 EI or decrease the range OR right click the Marker bar and select Edit V a ih UN ai Marker The Edit Marker box appears Adjust the Boundary field Mas Heterozygote Imbalance as x values as necessary Miri Heterozygote Imbalance Do ET z Additionally if a Marker needs only slight adjustment to the right RE HR et Al ake or left right click the Marker bar and select Adjust Marker The Ej Marker will move automatically to align with the closest peaks dc Uaa F N 2zx E Filtering Parameters M Min Homozygote Intensity Sets the minimum RFU value at which the software will call a peak if it is the only peak in a marker The number of peaks in a marker is determined by the number of i OF peaks above the Min Heterozygote Intensity level er Min Heterozygote Intensity Sets the minimum RFU value at which the software will call peaks if there is more than one peak in a marker The number of peaks in a marker is determined by the number of peaks above this minimum intensity level NOTE If the minimum homozygous and minimum heterozygous vaues are different from each other a single peak above the inconclusive heterozygous Min Heterozygote Intensity but below Min Homoz
237. the user to compare the same data set two different projects and detect differences based on a number of parameters including peak size and height quality score and commented alleles See Chapter 8 Additional Tools Pedigree File Name Match Allows the user to automatically add additional files to a previously created pedigree tree A smp file is exported See Chapter 8 Relationship Testing File Name Group Tool Used specifically with the MSI and LOH applications the Filename Group Tool allows users to define how reference samples and tumor samples should be grouped or paired A Text txt file is exported See Chapter 9 Additional Tools Convert Text to Binary Files For customers developing their own instrumentation the Convert Text to Binary Files option allows users to upload four or five color Text files without headers for conversion into SCF four color data or SG1 five color data trace files for analysis with GeneMarker HID See Chapter 9 Additional Tools Replicate Comparison Many labs choose to run and process multiple replicates of their samples This ensures that a genotype is still available in cases of contamination allele drop out or reaction failure Concordance between replicates can then be used to export the consensus genotype of the sample 33 August 2013 Chapter 3 Main Analysis Overview Output Trace Data Provides the option to output the raw or sized trace data as a TXT or SCF file Select the samples to
238. tifier Enter common filename nomenclature for Ladder samples in the dataset must be in all capital letters Upon first analysis GeneMarker HID will automatically scan the dataset filenames for the Ladder Identifier values and subsequently label the Ladder samples with an LD and display the sample filename in blue font in the Sample File Tree Defaultis LADDER NOTE The Ladder Identifier option affects the operation of the Auto Select Best Ladder and Automatic Panel Adjustment features in the Run Wizard Additional Settings box After modifying the Ladder Identifier field re activate Run Wizard and proceed through Data Process The Auto Select Best Ladder and Automatic Panel Adjustment features may now be selected Positive Control Identifier Enter common filename nomenclature for Positive Control samples in the dataset must be in all capital letters Upon first analysis GeneMarker HID will automatically scan the dataset filenames for the Positive Control Identifier values and subsequently label the Positive Control samples with a PC and display the sample filename in green font in the Sample File Tree Default is PC Negative Control Enter common filename nomenclature for Negative Control samples in the dataset must be in all capital letters Upon first analysis GeneMarker HID will automatically scan the dataset filenames for the Negative Control Identifier values and subsequently label the Negative Control samples with an NC and dis
239. ting Database Submit the genotypes from deconvoluted mixture sample Major contributor minor contributor or unknown The contributor label is unknown if the mixture approximately 1 1 neither file fits the definition of major or minor contributor Submit Major Submit Minor Submit Unknown Contributor Search Relationship Testing Database Search for exact matches and the likelihood ratio results without navigating to the Relationship Testing Application Results are displayed in the mixture analysis screen Search Unknown Contributor Confirm Report Options Unconfirm Right mouse click on the mixture analysis report table results to activate edit confirmation Confirm Locus submit copy and save options Unconfirm Locus Submit Major 9 9 EIE Submit Minor s neCanhbun Ha Make rcu Maer Ma esos Mac HIM Mee HIM dye tang Recon c Two Conte fan 1113 08 33 158 o Cornu 1 Cornetanat 2 LR E Lm 1 2 3 33 ir aces 8 X579 daan 1134 423 anii 135 3 098 ome O78 076 083 ames 07 nen 087 um 095 n5 09 qun 86 073 143 doo 144 0092 nas 10032 095 nmm 0 nas nomo as 0 0023 gassm 3 35 53470 Copy Export Gas iow s2 032 t 0064 93 0 0006 ag 0 0062 n4 ppm 0 75146 111215 Tomea 20 2224 Copy Damen ipn 035 0 U201 nas 00013 ngs 0003 ng 006 ns 1041 nal mes Save and Export Result Table
240. tion User Manager History Setting Overtime Protection eS A When selected GeneMarker HID will logout the user after the specified time entered in the Wait field When the user is logged out WV Record Data Edi Hoy the status of the analysis remains unchanged until the user logs back in with username and password Record Data Edit History w Run User Protection 130 August 2013 Chapter 10 User Management When selected any changes made to the allele calls of the project will be saved in the Edit History log Please see Edit History section below for more information Edit History Audit Trail When Record Data Edit History is selected in the User Manager Settings box see User Management Settings section above any change to allele calls in the analysis will be recorded Changes can also be recovered in the Edit History feature Procedure 1 Click the Show Chart Table icon in the Main Analysis window EEN 8 B EE BEA We we j j 2 The Peak Table will appear below the sample electropherogram 3 Make changes to allele calls by right clicking any cell in that allele s row in the Peak Table or right click the grey vertical bar at the center of the peak in the electropherogram 4 Choose to Edit Allele Edit Comments Add Delete Allele and Confirm See Chapter 3 Main Analysis Overview 5 Oncea change has been made to the allele call notice the pink shading in the No column of the Peak T
241. tion and Independent Assortment The law of segregation describes how phenotypes are an expression of two alleles inherited independently one from each parent Independent assortment is the idea that the inheritance of one gene or allele does not affect the likelihood of inheritance of a different gene Today we have found that this is not true of all genes some genes are invariably linked and inherited together Linked genes do not follow Mendelian inheritance patterns With GeneMarker HID s Pedigree module Mendelian inheritance patterns can be analyzed That is alleles are inherited independently one from each parent and alleles are not influence by the inheritance of other alleles Forensic STR loci were selected because they follow the laws of Mendelian genetics In the example below the child s Marker D75820 is highlighted red The child s Allele 8 matches with the mother and father however the child s Allele 10 does not match with either the father or the mother Because Allele 10 does not match an allele from the father or mother it can be assumed that one of the parents is not related to the child Because all three individuals share Allele 8 it cannot be determined which parent is not related to the child Review additional loci to determine which parent is unrelated 99 August 2013 Chapter 8 Relationship Testing Mendelian Inheritance of Alleles ill Pedigree Edit C SoftGenetics Forensics Frag Data DDC Paternity Data DDC
242. tool is applied in the main analysis window Icons and Functions l The following icons are available in the Print Preview window prior to printing the Print Report Export Formal PNG Image PNG Iiag File Namng M JPEG Image Print PDF file _ Opens the Print options box Select a printer the print range and r the number of copies Named by page number Save Group Samples as One File Export to File Export Directory Opens the Export Report to Files box Save each page of the Print C Program Faes WB6 SoltGenetics GeneMarker_ Report as an individual image file JPEG PNG or PDF Select the directory to export the files to When saving as a group the file name will be 2a the first file of the group See View Preferences Report Settings if a prefix of Ladders and Controls is desired for allelic ladder positive and negative control files Named by sample name saves each PNG or Name Type JPEG under the sample name JPEG Image n7 Dljpg JPEG Image A08 02 ipg JPEG Image 708 02 JPEG Image ue D5 jpg JPEG Image Start by Page Number combines the page number and the sample name for the pati cov n5 Oi ipg JPEG Image saved file name amp Pgl2 C08 06 jpg JPEG Image Mame Named b b h file by th b ithin th t amed by page number saves each file by the page number within the repor ic T uA GeneMarker Pgz ipg 7e GeneMarker
243. trace Single left click samples in the Sample List to see additional samples OR use the Up Down Arrow keys The green triangle peak indicators appear atop peaks that correspond to the enabled Sizes in the Size Standard Trace The basepair size associated with the green triangle peak indicator is located above the electropherogram The peaks selected for size calling can be edited in the Sample ILS frame as described below Navigation in the Sample ILS frame is similar to navigation options in the Main Analysis window See Chapter 3 Main Analysis Overview Editing Size Call Single left click a green triangle peak indicator to select it The triangle that is currently selected will be yellow To move the green triangle hold down the CTRL key and left click and drag it to the desired position Right click the green triangle peak indicator or right click the top of an unmarked peak to see additional options Delete Peak 45 August 2013 Chapter 4 Fragment Sizing Standards Removes the green triangle peak indicator from the Sample ILS and the peak will not be considered in the Match Score calculation The Match Score calculation is updated when Update Calibration is selected Add Peak Right click at the peak position and select Add Peak A green triangle peak indicator will appear at the cursor position To move the green triangle hold down the CTRL key and left click and drag it to the desired position The newly added peak will be included in
244. uals contributing to the mixture Consider all possible allele combinations VV VV V Y Compare References Samples GeneMarker HID identifies the presence of potential mixture samples designates allele peaks and calculates peak area or height ratios in the main analysis screen The Mixture Analysis Application is activated from the Applications menu in the main analysis screen Mixture analysis identifies the mixture samples and any single source contributor samples in a file name tree considers all possible allele combinations calculates the Mixture Ratio residual score heterozygous imbalance for each genotype combination and calculates the likelihood ratio for single source samples that are potential contributors to the mixture Often as in the case of male female cell separation a single major contributor profile of the perpetrator can be identified in the results table This genotype can be exported directly to the Relationship Testing database in GeneMarker to search for exact matches or close relatives even in cases where there is no suspect reference sample Procedure Identify the Presence of a Mixture The presence of a mixture is determined by setting the needed parameters in the third plate of the Run Wizard The criteria are used for identifying potential mixtures include presence of more than two peaks in for a marker severe peak height imbalance between alleles of a locus also see Panel Editor stutter peak filter chapter 5
245. units plotted along the x axis in the original Raw Data Analysis window are converted to basepair size units as defined by the Size Standard selected and the Internal Lane Standard ILS of the individual samples Fragment mobility is from right to left with the smallest size fragments on the far left of the trace The Peak Table contains information about the called peaks currently displayed in the Electropherogram Electropherogram Trace Display Range The basepair size range x axis is as set in the Run Wizard Data Process Allele Call options box The RFU range y axis is variable and will re adjust according to the maximum peak height in the trace To manually set x and y axis ranges use the Set Axis icon in the main toolbar Cursor Locator The x and y axis position of the mouse pointer in the electropherogram is displayed in the upper right corner of the electropherogram Allele Call If a Panel is applied to the data then grey horizontal bar Markers will appear above the electropherogram indicating locus ranges Bin ranges appear as dye colored brackets above and below the sample trace Allele Labels appear below the electropherogram and are associated with the center of each called peak which is also marked by a light grey vertical line in the electropherogram If a Panel is not applied then Allele Labels for called peaks will only indicate the basepair size of the peak Im 24 August 2013 Cha
246. us FU Uc Fax E aS 59 IMPORT PANELS FROM 56 IMPORT PRE DEFINED 5 59 IMPORT SIZE STANDARD ee eee eee eee ee ee eene sehe eee eene enun 42 INDEPENDENT ASSORTMENT cccccsccccscsccscscsccscsccscscsccscsccccecs 93 INDEPENDENT 99 INDIVIDUALAD sieve 94 113 INDIVIDUAL SAMPLE ACCORDANCE FILE eee eee een 96 INHERITANCE PATTERNS ccccccccscscsccscsccscccsccscsccscscsccscscsccscs 93 INSERT ALLELE icu RU Lu uH Du CRRKR 26 INSERT SIZE ge P 40 INSTALLATION dus dus oie ao 5 INSTALLATION WIZARD ula cuta va va va tn v ta an 5 6 8 18 INTENSITY COEFFICIENTS cccccccccccscscscccscccscsccccscscscscscscsccecs 16 INTERNAL LANE STANDARD ILS esses 39 K KINSHIPANALYSIS 516292015923 cocavacdeeaisssavensevessenseeanes 92 101 KINSHIP 101 L LABEL DYES 5 72 LADDER IDENTIFIER 2ccccccccccccccecscscscscscscscsccccscscscscscees 18 30 LADDER LABEL coscscsaudessosdntasseasabacansdasadinadededecaseiasteauseaas 30 LANE SCORE seve vaa de apes VENERE RUSSIE DE 44 LINE TIST Qut eue peque oue aa
247. ust 2013 Chapter 4 Fragment Sizing Standards sample s ILS peaks could not be aligned with the Size Standard selected then the sample will be marked with a red strike through When low score or failed samples occur select the Size Calibration Charts icon in the main toolbar to correct the size calling Low Match Score and Failed Samples fa e Xj di GeneMarker Untitled ie Project Applications DE 8 E S Allele Call Tools Hej n k 6 6 bs EE l v Marker None d Untitled 5 H A Raw Data 564 T Group1_0 07 fsa B 564 B Group1_0 05 fsa 50 100 554 B Groupl 102104 05 fsa 564 T Group1_0 07 fsa 564 T Group1_102104_07 fsa amp 745 B Group1_0 09 fsa B 745 B Groupl 102104 08 fsa B 745T Groupl 0 11 fsa B 745 T Groupl 102104 11 fsa 250 350 400 450 500 564 T Group1_102104_07 fsa x 50 100 150 200 250 350 400 450 500 Page Up Page Down Size Calibration Charts The Size Calibration Charts tool is designed to aid the user in determining success or failure of size call after GeneMarker HID s automatic sizing is performed Click the Size Calibration Charts icon in the main toolbar of the Main Analysis window As mentioned previously once a Size Standard has been applied to the dataset Size Match Score indicators appear next to the filename in the Main Analysis window Sample File Tree Samples with a high Match Score are indicated by a
248. vssnveveesuresvecssvesvsesvveste 97 F FAMILIAL RELATIONSHIP cccccccscsccscsccscscsccscscsccscsccscscsccscsess 93 vain 2609 ticks bene 2e 98 103 108 FAMILY NAME EUR FOU MN On M ME 94 113 9 112 3 tL 94 114 FILE LABELING CONVENTIONS ccccccscsccccscsccscscsccscsccscscsccscsoes 30 FILE NAME aon ko au au naa 118 33 118 122 FILE TYPES 11 FILENAME GROUP EDITOR ee ee ee eene eene eee eee eene neenon 118 FILTERING PARAMETERS ccccscscsccscsccccscsccscscsccscsccscscsccscsoss 53 FIX SIZE 46 FIXED BIN WIDTH 54 FORENSIC PREFERENCES 30 FORENSICS REPORT STYLE 2i 67 FRAGMENT MOBILITY anao ua 39 FRAME i eeu e Pepe te ee De aa qe veau ee vs 16 FREQUENCIES soson p coRp dps valebodipe sius ocn MAD PEE DERE 98 G GEL IMAGE ESPERE RENS REA MIA 60 GEL IMAGE DISPLAY 24 GEL IMAGE 5 5 5 30 94 113 GENE FREQUENCIES ccccccccccsccecscscscccscscsccccscscscscscscscsecscsces 98 GENEIMIARKER HID LOCAL 5 6 8 GENEMARKER HID NETWORK VERSI
249. w the data is processed and printed Add Samples to Project Run amp b Print Report Activates the Run Wizard and begins the data processing setup This allows the user to select or adjust program settings in a sequential manner The same process action can also be accomplished by clicking the Run icon in the toolbar Il Options En Project Comments Apply Sample Grouping 31 August 2013 Chapter 3 Main Analysis Overview Auto Run GeneMarker HID will process data using the last set of parameters selected If one or more of the parameters require changing to improve analysis select Project Options change the desired setting s and re process the samples for analysis Add Samples to Project The user can add samples to a project that has already been sized and analyzed When selected the Open Data Files box will appear Click Add to select individual files to the project and click OK The raw data file will be sized and processed with the same settings as the other files in the project and added to the bottom of the Sample File Tree Print Report Selecting Print Report launches the Print Report Settings box which allows the user to define display settings in the Print Report The software permits printing of the sample electropherograms You can choose to print all samples selected samples or print samples along with the allele table if desired See Chapter 6 Reports and Printing Options Allows you to access and change pa
250. would like to zoom in on To zoom back out hold down the left mouse button and drag a box in the opposite direction from lower right to upper left The user may also use the Zoom icons in the main toolbar to zoom in and out The main analysis window also allows the user to manually set the x and y axis with the Set Axis icon Horizontal Movement Ihe Synthetic Gel Image and the Electropherogram are synchronized to allow the user to view both images at once To move the images in the horizontal direction use the top slider bar below the toolbar to scroll the image in either direction or hold down the right mouse button and drag the trace right or left Marker Locus Specific Viewing To scroll through individual markers loci select a marker from the Marker drop down list in the main toolbar To view subsequent markers use the Up Down Arrow keys Synthetic Gel Image Features The Synthetic Gel Image displays all samples in the dataset vertically The direction of fragment mobility is horizontal with the small size fragments on the left and the larger fragments on the right so that the gel aligns with the electropherogram trace display Move the mouse pointer over the Synthetic Gel Image to reveal the sample lane filename Image Utilities Click the Image Utilities icon in the upper left corner of the Synthetic Gel Image A fly out menu appears with the following options Copy to Clipboard Copies the Synthetic Gel Image to the
251. xact match Output Trace Data Tools Output Trace Data The Output Trace Data tool exports raw or sized data of uploaded sample files as Text txt or SCF scf files Procedure 1 Select whether to export the data as a Text or SCF file 2 Choose the directory and folder to save the exported data to in the Output File Name field 3 Select the samples to include in the output file from the Select Samples field Select which dye color data to export from the Select Dyes field Select whether to export raw or sized data from the Data Type options 6 Click Export to export the data to the specified folder Project Comparison a 119 August 2013 Chapter 9 Additional Tools Tools Project Comparison The Project Comparison tool can serve three functions First it can be used to compare two independent analysts analyses Second it can be used as a validation tool to determine differences in allele calls based on analysis parameters or instrument runs Third it can be used to compare the projects if the sample names are not identical as would be the case if the samples were analyzed on two different genetic analyzers Procedure for comparison of projects with same file names 1 After initial dataset analysis select Tools Project Comparison 2 The Project Comparison window appears 3 Click the Open Project to Compare icon 4 Use the file directory window to locate and select a previously saved So
252. ygote Intensity will be called and labeled with a Check Quality and LO Quality Reason if a second peak is detected above the N x Stutter Filter value The second peak will not be called however because it is below the Min Heterozygote Intensity threshold Inconclusive Range If desired set an inconclusive range for homozygous and heterozygous peaks Peaks within the inconclusive range will be flagged with a Check Quality and IHO inconclusive homozygous or IHE inconclusive heterozygous Quality Reason This setting is helpful for flagging peaks that are above the minimum detection level but are not high enough to include in statistics within the stochastic range Max Heterozygote Imbalance Uses the percentage of the highest peak in a marker to define the maximum peak threshold For example if the threshold is set to 60 the height of all allele peaks must reach at least 60 of the height of the highest peak in that particular locus If a peak does not reach that height it is flagged with a Check Quality and IMB Quality Reason Min Heterozygote Imbalance Uses a percentage of the highest peak in a marker to define the minimum peak threshold If a peak does not reach the minimum imbalance threshold the peak will not be called This Apply Stutter Settings to All Markers 53 August 2013 Chapter 5 Panel Editor function is the equivalent to a filter allowing users to filter out peaks that are less than a given percent of the highest pea

GeneMarker® HID User Manual

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