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KaryoStudio v1.2 User Guide

1. 7 IA ECCE TET RET ETET TOT LL TR TT TI 8 Creating a New Project 8 Loading Sample Intensity Data Using a Sample Sheet 14 Loading Sample Intensity Data by Selecting Directories 21 Opening an Existing Project 30 Assessing the Quality of Your Data 30 User Interface 33 eT OOUGTION e 40 15 cannes TTE nae oda a SR la a 34 Main Window Menus 35 Table Window 37 Found Regions Table 37 Known Regions Table 39 viii 1 Chapter 4 Chapter 5 Database of Genomic Variants 42 Samples Table 44 Chromosome Browser 45 PUNCHOM kiaina ssQESEESREPSS IURE 46 Navigating the Chromosome Browser 47 o C rrr 52 Duo and TrioView 56 Log WINGOW 4 4 254508 Hee oe diarenn de anta es su aa 57 Analyzing Found Regions 61 Laias e Tei o cree REDLIAAP ER osas canso pata eo oe ee 62 Filtering Found REGIONS etico toi ee eee aes 63 Displaying Found Regions 65 Scrolling Through Found Regions 67 Sorting Found REGIONS uad we es 6 ees sus ee ray Ea 68 Adding or Editing Comments
2. 70 Adjusting Found Region Parameters 70 Editing Confidence Score or Estimated Copy Number 70 Editing Start and Stop Positions 73 Exporting Found Regions 82 Cross Matching to Known Variants 85 MoCU TORN e iei 5 aio te ips ed a T 86 Database of Genomic Variants DGV 87 Updating KaryoStudio with the Latest DGV Build 87 Using the DGV with KaryoStudio 87 DOY DAS Ane aaa any er ed aia ai 94 Updating the DGV Table and Track 96 OMIM Online Mendelian Inheritance of Man 96 Using OMIM with KaryoStudio 96 UCSC Genome Browser 102 Using the UCSC Genome Browser with KaryoStudio 103 PUDM cu 104 Using PubMed with KaryoStudio 104 BIO una RN sore desa da eae oe Re eh bid 104 Using DECIPHER with KaryoStudio 105 o a AI KI AI ee A R ee EA 106 Using Ensembl with KaryoStudio 107 CHOP Database MP 108 Using the CHOP CNV Database with KaryoStudio 109 Part 11328837 Rev B Chapter 6 Appendix A Appendix B Appendix C Appendix D KaryoStudio v1 2 User Guide Table of Contents Generating a Cytogenetics Report 111 Ine e 112 What s in a Cytogenetics Report 112 Adjusting the Information in a Cytog
3. Export data ERU OE DUM Select Set of features to render select SS Cavia porcellus Monodelphis domestica Output format HTML Omnia md e mem m mo cmd 1 Figure 88 Ensembl CHOP Database The Copy Number Variation project at the Children s Hospital of Philadelphia CHOP represents an effort to identify all frequent copy number variations CNVs that exist in the human genome The database currently consists of data from over 2 000 healthy individuals This database was originally described in the following manuscript TH Shaikh et al 2009 High resolution mapping and analysis of copy number variations in the human genome A data resource for clinical and research applications Genome Res 19 1682 1690 doi 10 1101 gr 083501 108 Part 11328837 Rev B CHOP Database 109 Using the To use the CHOP Database with KaryoStudio do the following CHOP CNV Right click a found region and select CHOP Database from Database with the context menu Figure 89 KaryoStudio ff Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin V2 4 4 0 C Docum O Uu d Ic QoS Gis EB T 1 5x 1 2x Found Regions Known Regions DB of Genomic Variants Ww WP Uncheck All Value Cor 14 N 12134_R 1 147305744 147467750 162006 30 Database of Genomic Variants Browser 17 Database of Genomic Variants Defects OMIM UCSC Genome Browser PubMed DECIPHER ENSEMBL q CHOP Database Samples Copy row to Cli
4. 247200 and the Wolf Hirschhorn syndrome 194190 are other examples of terminal chromosome deletions Y Nakamura et al 1997 found that disruption of the S pombe telomerase gene resulted in telomere shortening and senescence and expression of mRNA from the human gene correlated with telomerase activity in cell lines Meyerson et al 1997 found that the human EST2 transcript was upregulated concomitant with activation of telomerase during immortalization of cultured cells and was downregulated during in vitro cellular differentiation These observations suggested that induction of EST2 mRNA expression is required for the telomerase activation that occurs during cellular immortalization and tumor progression Edit History Gene map Activation of telomerase the enzyme that synthesizes the telomere ends of linear chromosomes has been implicated in human cell REESE immortalization and cancer cell pathogenesis Enzyme activity is undetectable in most normal cells and tissues but present in immortal HA ko cells and cancer tissues K olquist et al 1998 used in situ hybridization to study TERT expression at the single cell level in normal TENE tissues and in various stages of tumor progression In normal tissues including some known to be telomerase negative TERT mRNA was present in specific subsets of cells thought to have long term proliferative capacity This included mitotically inactive breast lobular LinkOut epithelium in addition to s
5. KaryoStuaio v1 2 User Guide Software for Cytogenetics Data Analysis and Reporting FOR RESEARCH USE ONLY ILLUMINA PROPRIETARY u m n a Part 11328837 Rev B KaryoStudio v1 2 User Guide Notice This document and its contents are proprietary to Illumina Inc and its affiliates Illumina and are intended solely for the contractual use of its customer in connection with the use of the product s described herein and for no other purpose This document and its contents shall not be used or distributed for any other purpose and or otherwise communicated disclosed or reproduced in any way whatsoever without the prior written consent of Illumina Illumina does not convey any license under its patent trademark copyright or common law rights nor similar rights of any third parties by this document The instructions in this document must be strictly and explicitly followed by qualitied and properly trained personnel in order to ensure the proper and safe use of the product s described herein All of the contents of this document must be fully read and understood prior to using such product s FAILURE TO COMPLETELY READ AND EXPLICITLY FOLLOW ALL OF THE INSTRUCTIONS CONTAINED HEREIN MAY RESULT IN DAMAGE TO THE PRODUCT S INJURY TO PERSONS INCLUDING TO USERS OR OTHERS AND DAMAGE TO OTHER PROPERTY ILLUMINA DOES NOT ASSUME ANY LIABILITY ARISING OUT OF THE IMPROPER USE OF THE PRODUCT S DESCRIBED HEREIN INCLUDING PA
6. Maco M eo co cn uo 299207 SOLLL WN BLES UTN 299207 SOLLLWN BLES LN 299Z0 7N SOLLLWN BLES LN 299Z0 7N SOLLLWN BLESL VN 299Z0 7N SOLLL WN BLES WN 299Z0 7N SOLLL WN BLES LN 299Z0 7N S91LL LIN BLES UTN 29920 7N S9LL LIN BLES UTN 299Z0 7N S9LL VvN BLES UTN 299Z0 7N SOLLLWN BLES LN 299Z0 7N Sg V VN BLES LN Figure 36 Karyotype View Window Three Samples Selected 3 To display information about a found region in the info bar at the top of the window hover over a found region The information displayed includes sample ID chromosome start and end coordinates and copy number value Figure 37 Part 11328837 Rev B Karyotype View 55 NA18319 NAI1165 N402668 NA18319 NA11165 N402668 NA amp 16319 NA11165 NAO2666 NA16319 NA11165 NAO2668 N418319 N411165 NA18319 N402668 NA11165 NA02668 NA16319 NA11165 NA18318 NAD2B6S NA11165 NAO2658 Found Region Information Click a Found Region NA18313 NA11165 NA402668 NA18319 N411165 NAO2668 NA16319 NA11165 N402668 NA16319 NA11185 NAO2658 NA18318 NA11165 NAO2668 NA16319 NA11165 NA02658 NA18319 N411165 N 402668 w Found Region Information To display a region in the chromosome browser click a found region Figure 38 Found Region in Karyotype View Window NA18319 NA11165 N402668 Karyotype View Found Region N418319 N411165 N402668 4 NA18319 NA11165 N402668 N418319 N411165 N402668 N
7. Figure 47 FoundRegionsFilterForm The Known Regions File to be used in the project is listed in the second text field To change the Known Regions File click Load and browse to the file you would like to use instead See Appendix C of this document for information about formatting a Known Regions File Different classes of found regions can be assigned different filter settings e Settings for regions that at least partially overlap with regions defined in the selected Known Regions File are labeled Inside in the InKnownRegions column e Settings for regions that do not overlap with regions defined in the selected Known Regions File are labeled Outside in the InKnownRegions column e Settings for Gains Losses and Copy Neutral events CNLOH can be set independently in the column TypeOfCNV Optional You can define found regions from which data should never be displayed by selecting the Use Ignore Regions File checkbox and loading a file Part 11328837 Rev B Displaying Found Regions 65 In order to be ignored a Found Region must fall completely within an Ignored Region This feature has many potential uses such as ignoring common polymorphic CNVs or constraining analysis to a portion of the genome For example you could ignore all chromosomes except 21 to analyze only aberrations on chromosome 21 See Appendix D of this document for more information about formatting an Ignored Regions File 4 To save the sett
8. Loading Sample Intensities KaryoStudio Project 22 Please specify the samples you want to load by identifying the U m n a sample sheet and associated data and manifest repositories Sample Sheet C Documents and Settings aleoniDesktop eS_ v1 2 Project_Info Hu Data Repository Browse Manifest Repository Browse Cancel lt Back Mext gt Finish Figure 10 Wizard Sample Sheet 16 CHAPTER 2 Loading Your Data 3 Browse to the repository that contains the data you want to include in this project Figure 19 You must specify a path in the wizard E NOTE However if a different path is specified in the sample sheet the path in the sample sheet overrides the path you provide in the wizard KaryoStudio Project Wizard Loading Sample Intensities KaryoStudio Project www Please specify the samples vou want to load by identifying the ul m n a sample sheet and associated data and manifest repositories Sample Sheet gs and Settings aleon DesktopikS _v1 2 Project_Info Hui Browse Data Repository ges and Settingsaleon Desktoples v1 ziPrajeckE InFolIDz Browse Manifest Repository Browse Cancel lt Back Next gt Emish Figure 11 Wizard Data Repository Part 11328837 Rev B KaryoStudio v1 2 User Guide Creating a New Project 17 4 Browse to the repository that contains the SNP manifest you want to use
9. Pia Gariri P A copy number wwii using DAL hd ret 2007 ERES ony number wa variation rupi desir WAA Hum Cer O Cercs a mee Ced Ey over 109 Publi Cantrell rticios t A mri OOH an patiti Vance c fyndnone M end reve A Med Gabel US amp Copy number variation in the jog cire Dads 20098 b The piisa ar paham comparison iple cop iur mnimy leche a ERAS az t Xem d Canada If you need more information about this database go to http projects tcag ca or send email to dgv contact sickkids ca DGV Data All of the CNVs in the DGV were identified in healthy control samples The DGV is constantly being updated as new studies deposit information and as technology evolves For the latest build and latest information please see the DGV web page http projects tcag ca variation When you click the DGV tab it may take a moment to load When it does scroll to the bottom of the list Figure 77 Part 11328837 Rev B Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin V2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 122 testC DUA LAAk EM 1 sx 1 2x 2x SN 4 D N Found Regions Known Regions OB of Genomic Variants Database of Genomic Variants DGV index ID Landmark Ch Stat Variation 37350 Warnation_37 381 Variation 3302 Wanation_ 37383 Variation 3 384 Variation 37385 Variation 37 38b Variation 3 387 Variation 37388 V ariatior 37383 Variation 37330
10. S E E E E Main Window Menus 35 Table 1 lists toolbar button options available from KaryoStudio s main window KaryoStudio Toolbar Button Options Name Create New Project Open an Existing Project Save the Current Project Close the Current Project Write Reports to Project Directory Close Application Expand to Chromosome Previous Chromosome Select Chromosome Next Chromosome Pan Up 1 4 Step KaryoStudio v1 2 User Guide Function Create a new project Open a previously generated KaryoStudio project Save the current KaryoStudio project to your computer Close the current KaryoStudio project Generate a paf report and save it to your computer Close KaryoStudio Expands the current Chromosome Viewer view from selected found region to entire chromosome Display previous chromosome in the Chromosome Viewer Select chromosome to display in the Chromosome Viewer Display next chromosome in the Chromosome Viewer Adjust view slightly towards the beginning of the chromosome 36 CHAPTER 3 User Interface Table 1 Toolbar Button s E E EE 9 E 8 s Ee p iiy Es KaryoStudio Toolbar Button Options Name Pan Down 1 4 Step Zoom in 5X Zoom in 2X Zoom out 2X Zoom out 5X First Aberration Previous Aberration Next Aberration Last Aberration Settings Display Log Function Adjust view slightly towards the end of the chromosome Adjust
11. This brings up a list of scientific publications in which the key word has been found Note that the more common your inquiry the more hits you will get To reduce the number of hits make your search term as specific as possible The DECIPHER consortium is a network of clinical genetic centers KaryoStudio allows you to examine a specific found region of interest against this cytogenetics database of clinical cytogenetics cases by right clicking on the found region and sending it to the database If a particular portion or the whole region of your found region has been previously linked to a certain phenotype you can view that here From the DECIPHER database you can view your found region within the Ensembl Genome Browser identify the type of array or technology used to identify the specific region and obtain a DECIPHER Syndrome Report for a specific phenotype of interest including a clinical description references affected genes and a phenotype report Part 11328837 Rev B DECIPHER 105 Using To use DECIPHER with KaryoStudio do the following DECIPHER gt Right click a found region and select DECIPHER from the with context menu Figure 85 Ka ryoStudio Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Docume id 1 5 1 2x DOA Bl c4 QD E Gi Found Regions Known Regions DB of Genomic Variants XY Uncheck All Value Cor 14 N412134 AR 1 147305744 147457750 162006 1 38 mi Ni
12. 3 N 1 n PPM HA MPH MP 7 NATE Database of Genomic Variants Browser 48E NATE Database of Genomic Variants Defects 47 NATE 56 4 NAME UCSC Genome Browser 49 2 NAE PubMed DECIPHER gt ENSEMBL CHOP Database 0 0 C ro Samples Crid Copy row to Clipboard Deve Copy all to Clipboard Wolf Prad Copy DBGY CHR Format Prad Mille Edit a Found Region Retinoblastoma NAUTZBSU WIBUUSJJTZE DNA DO Turner Syndrome NA02668 W G0033128 DNA Fa b M 533 puo J EH S AA IA 1 HN LHOD HM ZZ94NZ sHvMG M Tawa zA2 v zStozrqi4 4 orm H peS690L00 WN dsva HWwNv H zawwio HH cawisvd vSols I PLEEL DOT 1 SALA I aso I 199 IL l TS Was HE ZOSZOLLOO WN d LYZSivl 4 zsiozri4 esor j 2 00 1 000 00 1 00 2 00 Smoothed Log F CO MN CH CO PO The project has been loaded successfully Sample Cri du Chat CHR 5 7614901 22322881 Figure 61 Editing a Found Region 7 Select Edit a Found Region from the context menu The FormSettings screen appears Figure 62 KaryoStudio v1 2 User Guide 80 CHAPTER 4 Analyzing Found Regions FormSettings El Editable Comment Contidence Enabled Start Stop Value O Misc Chr Cytoband CytobandLocus Filtered ut GainOrLoss Genes ame Image Length Loci 5918 188 True 7614901 22322881 1 F p15 31 p15 2 p15 1 p14 3 pl5 31 p14 3 False Loss ADCY2 NM 0010895854 FASTEDS A none 14707380 luna Interface Locus F
13. APPENDIX C Known Regions File Introduction Table 13 lists and describes the required columns of a Known Regions File for use with KaryoStudio Table 13 Known Regions File Column Descriptions Column Region Disorder Chr Start End Length Length minus overlap Description ID code for the region Text description of the significance of the region Chromosome number of the region Start coordinate of the region End coordinate of the region Length of the region in base pairs Harper to provide info The figure below is an example Known Regions File Known Regions File Default Known Regions Table txt Notepad File Edit Format Regions Disorder Cyt ogenl CytoGena CytoGens Cyt OGe nd Cy toGen CytoGen Ct Gern CytoGen CytoGena Cy toGend cytoGendi Cytocenle Cyt oGenls cytoGenla Figure 93 View Help chr start End Length Length minus overlap 1936 microdeletion 1 o 27800000 27800000 Bartter3 classic 1 16100000 20300000 4 Bartter4 infantile with sensorineural deafness 1 1 NFIA Haploinsufficienc S0590 BS7 00000 lp pericentromeric region l 11 600000 120 700000 1q21 1 Microdeletion with d For thrombocytopenia abs lq pericentromeric region 14 2400000 153300000 short stature pituitary and aoa defects small sella t wan der woude 1 205300000 209500000 4200000 4 1q41 942 microdeletionsFryns 1 212100000 222100000 lq subtelomeric region 1 234600000 247249719 1 144 1 241700000 24724807
14. B Allele Frequency and Log R Ratio 121 The Log R Ratio is based on normalized intensity data and for a sample is the log base 2 ratio of the normalized R value for the marker divided by the expected normalized R value For loci included in GenomeStudio statistics such as Call Rate the expected R value is computed by linear interpolation of the R value at the SNP s theta value for a sample relative to the R values of the surrounding clusters Because no clusters are generated for loci in the Intensity Only category the Log R Ratio for these loci is adjusted so that the expected R value is based on the weighted mean of the cluster itself Log R Ratio is displayed the same way for these loci as it is for loci included in GenomeStudio statistics in tools such as the IGV Both SNPs and intensity only loci such as nonpolymorphic probes which usually have the identifier cnv in their names are displayed in the log R ratio plots in KaryoStudio For example if for a given sample and SNP with A theta value of 0 2 an AA cluster at theta 0 1 R 1 5 an AB cluster at theta 0 4 R 2 5 The estimated R at theta for the sample is 0 2 is 1 5 0 2 0 1 2 5 1 5 0 4 0 1 2 1 83 If the R value for the SNP is 1 6 the Log R Ratio is log2 1 6 1 83 0 196 cnvPartition automatically scans both the B allele frequency and log R ratio data for the presence of aberrations In regions of the genome with two copies the B alle
15. CO CO ro 0d GC Samples Cri du Chat NA16583 WG0083128 DN Developmental Delay NA08585 W G0083128 DN Wolf Hirschhorn Syndrome N400343 WGO0093128 DNA PraderWili_rep NA12134 R WGO0093128 DNA Prader Willi NA12134 WG0093128 DNA Mille Dieker Syndrome NAOS208 WG0093128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WG0093128 DNA Smoothed Log F Turner Syndrome NA02668 W G0083128 DN B pau pun y EH B BIBI 1 ii GN was MH czedwz SHvNa B JAWO 1 ZADOW zstozrqi4 4 orm H t25690L00 WN ldsv HMNv M ZONNLO HE come vs 1 Koka SPEI asolWsv3 I c9scoLLOO WN A zstozrid d TiNdOs 1 The project has been loaded successfully Sample Cri du Chat CHR 5 7614901 22322881 Figure 55 Selecting a Found Region of Interest KaryoStudio displays your found region within the chromosome browser 2 To expand the view so that you can see additional data adjacent to the beginning of the found region click Pan Up Figure 56 Part 11328837 Rev B Adjusting Found Region Parameters 75 Ilumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_vy1 21v122 testC gii OX E t 1 5x 1 2x 2x sx K 4 OD Found Regions i i i 9 Known Regions DB of Genomic Variants Cri du Chat Y Y Uncheck All B Allele Freq Index Sample ID Chr Stet stp Lengh Value Con 90 02505 07510 N416593 105587447 105849470 262023 NA16533 64316996 64813520 502524 N
16. Cytoband OS PTT 48 KaryoStudio v1 2 User Guide xil List of Figures Figure 33 Figure 34 Figure 35 Figure 36 Figure 37 Figure 38 Figure 39 Figure 40 Figure 41 Figure 42 Figure 43 Figure 44 Figure 45 Figure 46 Figure 47 Figure 48 Figure 49 Figure 50 Figure 51 Figure 52 Figure 53 Figure 54 Figure 55 Figure 56 Figure 57 Figure 58 Figure 59 Figure 60 Figure 61 Figure 62 Figure 63 Figure 64 Figure 65 Figure 66 Figure 67 Figure 68 Figure 69 Figure 70 Figure 71 Figure 72 Figure 73 Figure 74 DEN AAA 49 Chromosome Browser Alignment Cursor 50 Karyotype View Window One Sample Selected 53 Karyotype View Window Three Samples Selected 54 Karyotype View Found Region Information 55 Found Region in Karyotype View Window 55 Found Region in Chromosome Browser 56 Selecting Additional Data Plot to Display 57 Chromosome Browser Duo View 57 Chromosome Browser Trio View 58 Trio View Additional Plot Settings Selected 58 Single Sample View with Gene Display 59 LOG WANCOW Mur 59 Found Regions Table 62 FoundRegionsFilterForm 64 Found Region Displayed in Chromosome Browser 66 Found Region Displayed in Context of Whole Chromosome 67 Aberration Length Sorted Short to
17. If you do not have KaryoStudio v1 0 installed on your computer you can immediately install KaryoStudio v1 2 from the CD If you are a KaryoStudio v1 0 user you must uninstall this version from your computer before you can install KaryoStudio v1 2 If you do not uninstall KaryoStudio v1 0 before attempting to uninstall KaryoStudio v1 2 you will be prompted to do so You may also need to uninstall the cnvPartition algorithm you are currently using cnvPartition v2 4 4 will be installed automatically with KaryoStudio v1 2 To install KaryoStudio perform the following steps 1 Put the KaryoStudio CD into your CD drive If the Illumina KaryoStudio Installation screen appears continue to Step 2 If the CD does not load automatically double click the KaryoStudio lt version gt exe icon in the KaryoStudio folder on the CD i The CD includes two versions of 7 NOTE KaryoStudio l If you are installing KaryoStudio on a 32 bit computer click KaryoStudio exe If you are installing KaryoStudio on a 64 bit computer click KaryoStudio64 exe KaryoStudio v1 2 User Guide 4 CHAPTER 1 Overview The KaryoStudio application unzips and the KaryoStudio Ksi application icon ksj appears on your desktop Karvostodio Double click the KaryoStudio icon on your desktop If this is a new installation the Registration screen appears Figure 1 B Registration To better serve customers Illumina Inc i requesting bo regi
18. M As 16 NAT Database of Genomic Variants Browser 17 Database of Genomic Variants Defects OMIM UCSC Genome Browser PubMed DECIPHER ENSEMBL 4 Samples Cri d Dew tol Prac oo oon ee to Prader Willi Miller Dieker Syndrome Retinoblastoma Turner Syndrome CHOP Database Copy row to Clipboard Copy all to Clipboard Copy DBGY CHR Format Edit a Found Region NAT 2134 803208 804250 4602668 4600951 20 04 4600931 26 004 4600951 28 DNA Figure 85 Selecting DECIPHER KaryoStudio automatically sends information about the found region to DECIPER and opens the DECIPHER window for that region Figure 86 KaryoStudio v1 2 User Guide 106 CHAPTER 5 Cross Matching to Known Variants Logged in as public Centres Studies Array Types Syndromes Search 15 18742374 20063682 search eg Mental retardation developmental delay 17p11 2 17 38199474 40407206 Patient number Chromosome Start Position bp End Position bp Arraytype Patient View Ensembl View 1353 15 1 Patient View e cytoview 29111655 Spectral Genomics 1Mb Array MENTIR el cytoview 18461853 20224003 WTSI TPA Patient View 19050502 20224003 WTSI TPA Patient View 18419709 18804305 WTSI TPA Patient View 18590230 20278373 WTSI TPA Patient View 18747225 20285790 WTSI TPA Patient View 19108124 26199055 Agilent Human 4x44K Patient View 19109124 20636478 Patient View 18668297 20060120 Agilent Human 105A Patient View 19109124
19. NA16593 2 89665111 NA16593 91902273 NA16593 32349805 10 CO JO PM 000 4 Samples Cri du Chat Developmental Delay Wolf Hirschhorn Syndrome Prader Will_rep Prader Willi Miller Dieker Syndi ne Retinoblastoma Turner Syndrome a NA16533 1 147305744 NA16533 103148241 NA16593 144317436 144981571 64819520 80668665 22322881 89885025 92863522 32752830 N 16533 NA08585 NA00343 NA 12134 R NA12134 NA 3208 NA04250 NA 2668 6 NA 16533 105587447 105843470 262003 1 502524 2691359 14707978 219914 147467750 162006 413085 103174645 26404 1 1 1 1 1 961249 3 1 3 1 64135 WG0093128 DN4 A WG0093128 DNA w G0033128 DNA w G0033128 DNA w G0033128 DNA WGO00931 28 DNA WG00931 28 DNA4 WG00931 28 DNA 447 Completed report generation Sample Cri du Chat CHR 5 7614903 22322881 Samples Table EJ f 1 5x 1 2x 2x 5X N 4 5 H Cri du Chat B Allele Freq 00 025 05 0751 0 2 00 1 000 00 1 00 2 00 Smoothed Log F Figure 25 KaryoStudio Screen Configuration A ay puo Chromosome Browser 1 sv d eA l PSS630100 WN ZONNLO HH c EOS TEMO ZADIA SHLHELgO7 1 Sov viy TiNdowy 4 6S0 TI SHvNG B zsto0zrTa4 4 omi H H WSOLM4 I asome 27x84 Hi B HMNYv H 2952054400 WN ZZIANZ 4 cS lds 7S OZTA M OLOAW H c2 HH Gene Information Part 11328837 Rev B Main Window Menus Table 1 Toolbar Button P E 8 B9 9
20. Ochs Dialer d Manifasta A HumantyloSNP 131 0_11316975_4 Sampla N Rapicaa Paranti Paranta Path Rafaranta Raferanr 4 51 408 R TCUT WgcO SOT ADI Female NAIGESI Nalpmretssdatall serecas mage datawweggtic alli 0 451E4D0 RO2CDI WIGODS TZ B01 Male HADITS43 Wilmre amp sdataDlNserices image datswegghcall20OB 1029 4518409 ROSCOT coe CO Male MAIZT MAI2134 Rf Wilmrrfsdatall serices_image_dataweggticalh 0081023 450148 AAC WSUS DUET Female MAD Nulriri Biesdab aT serait eria dal gica lA Td da EA RSC Vers E EXE EN Male PT 7 The Wimirfedatal sanitas image datate aeaa 17 M410319 4 516409 ROGCOT WO POT Female MAITOSIS Wilmre amp sdata l serecas mage dataweggrc alleiona1023 B MADAE32 4518409 ROMS WIGODO T2 DI Female MAGES Wilmre amp sdataDlNserices image datswegghcall2DOB 1029 4 516109 RO2CD2 WICODSGIZ HO Male MADITA27 Wilmr amp sdataDMserices image datsweggreallannermaa 4 Ea ASC VAEN ALE hale MALHEHA Yaaa sdb ara se erae dal gica lA Td ASTER RCO Mc EX BEA Mala Bai HIH Wimirfedatal eericeas image datate aeaa 22 MADAWA 4 51E 09 ROSCOZ WOOO C02 Male NACE 200 WaltiretsdatadTsearnicas_enage_datalwggt wamzuia 3 MADDEEB 4518409 ROSCO WIODS TZ DO Linknown MAD2668 Viimrr sdatadlMeerices_image_datswggfealh2008 1029 24 Figure 92 Example Sample Sheet Part 11328837 Rev B 126 126 KaryoStudio v1 2 User Guide Appendix C Known Regions File Topics Introduction Known Regions File
21. a recent version of Adobe mw NOTE Reader to view pdf reports Adobe Reader is available free from get adobe com reader What s in a Cytogenetics Report A Cytogenetics Report contains all of the essential information required to quickly summarize aberrations found within a specific sample Within each report is an entry for each found region including an image of the aberration its size an estimate of the copy number the confidence value relative score and a list of the disorders that have already been associated with that specific region Figure 91 shows an example Cytogenetics Report Table 9 lists the elements of a report and identifies the elements that are adjustable Part 11328837 Rev B What s in a Cytogenetics Report 113 Sample amp Product Cytogenetics Report Algorithm Version Sample Cornelia de Lange A Build 36 1 a Manitest Humanc ytoSNP 12 2 1_ 4 bpm cnvPartition 2 4 4 Detected Region nes B Allele Freq CHR 00 025 05 O75 1 0 Comment Hay punoj A 525uwouy Mb 144 89 145 06 Locus q21 1 Start 144967972 End 1464531 47 Length 1485175 User Entered Value 3 146 04 146 2 Comment GA Gain Ep 146 36 1286 563 i 145 53 00 1 00 0 00 1 00 2 00 oihan Chromosome View Found Region Known Regions Known Regions 1421 1 Microdeletion with susceptibility for CytoGen6 142400000 148000000 Len 5600000 thrombocytopenia absent radius TAR 19 pericentromeric region CytoGen
22. characterized by well defined and distinctive phenotypic features and natural history Itis caused by triplicate state trisomy of all or a critical portion of chromosome 21 CLINICAL FEATURES OMIM Down System Information Page pal Information such as a description of the condition clinical features and more is provided UCSC Genome Browser The UCSC Genome Browser is a popular tool that allows you to examine gene information expression data and many other parameters within your found region Yo can also upload custom data tracks to display in the UCSC Genome Browser Part 11328837 Rev B Using the UCSC Genome Browser with KaryoStudio KaryoStudio v1 2 User Guide UCSC Genome Browser 103 You can use KaryoStudio to see what other genomic features sit within a found region All analyses are based on chromosome and position The UCSC Genome Browser shows you information about RefSeq genes mRNAs ESTs genes in other organisms There are also other options within the page that allow you to add or remove additional data tracks of information y The UCSC Genome Browser may take w NOTE awhile to load Be patient while your data is loading it will eventually display At the top the chromosome and position are identified Next information about RefSeq genes is listed Next there is mRNA and EST information which is useful for looking at locations of specific gene transcripts Next the c
23. identified and deposited in the KaryoStudio DGV Based upon chromosome and position the DGV shows you information about what is known about the method used to identify the region the reference including details of the study and a graphical reference to any CNVs found in the region KaryoStudio v1 2 User Guide CHAPTER 5 Cross Matching to Known Variants Perform the following steps to use the DGV with KaryoStudio 1 In the Found Regions table select the region you want to search for in the DGV Figure 66 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 122 testC DUHA ka s Et 1 5x sex zx N 4 gt DW Found Regions i i i g Known Regions DB of Genomic Variants Cri du Chat Y Y Uncheck All B Allele Freq Index Sample ID Chr Stat Stop Length Value Con 90 92505 07510 1 NA16593 91902273 92863522 961249 NA16593 1 147305744 147467750 162006 NA16533 2 89665111 83885025 219914 N 16533 5 7614901 22322881 14707980 NA16593 7 64316996 64819520 502524 NA16593 14 77377306 80665665 2691359 NA16593 14 105587447 105849470 262023 NA16533 16 32349805 32762890 413085 NA16533 x 103148241 103174645 26404 NA16593 x 144917436 144381571 64135 2BSOwvIM I TiNdow 4 10 0 MAA WMH c H TSOYA asoiWsv 7 mas Hi SZOLLOO WN zstozrid Samples 94N coena d OLOAW dH zz lisa HMNv H Cri du Chat NA16583 WG0093128 DNA Developmental
24. or a region associated with a condition For this reason KaryoStudio allows you to check a found region against a list of known regions as well as several different external sources This function allows you to determine which regions found in your samples may be the most relevant This chapter describes how to link to these external information sources from within KaryoStudio and how to use this additional information in your study KaryoStudio allows you to link to several databases including DGV Database of Genomic Variants to determine whether your region has been identified as a copy number variant in normal healthy individuals gt OMIM On line Mendelian Inheritance of Man to allow further mining of a particular phenotype region or gene of interest gt UCSC Genome Browser to view genes or numerous other information tracks for a particular region in this popular public genome browser PubMed to research publications relevant to a particular phenotype gene of interest DECIPHER to compare found regions to the DECIPHER database of submicroscopic chromosomal imbalances Ensembl to view a found region in the graphical Ensembl genome viewer CHOP Database to determine whether a found region overlaps with copy number variations described in this database of samples from healthy individuals Part 11328837 Rev B Database of Genomic Variants DGV 87 Database of Genomic Variants DGV Since the wi
25. pairs Copy number of the found region Gain or loss Confidence score for the found region calculated by the algorithm Part 11328837 Rev B What s in a Cytogenetics Report 115 Table 10 Cytogenetics Report Detected Region Information Column Data plot Regions tracks Coordinate ruler Comment Known regions Table 11 Cytogenetics Report Footer Column Sample Name Manifest Signature and date Username KaryoStudio v1 2 User Guide Description Plot of B Allele Freq and Log R of markers in the found region Found Region Known Region and DGV tracks from the chromosome browser Chromosomal coordinates of the data plot in megabases User defined comments for the found region Can be left blank Regions known to overlap with the found region Includes Region Name Region ID Chromosome Start coordinate End coordinate and Length Description Name of the sample presented in the report Product manifest used to create the project Area for the investigator to sign and date the report Name of the user logged into the system when the report was generated 116 CHAPTER 6 Generating a Cytogenetics Report Adjusting the Information in a Cytogenetics Report Generating a You can adjust the information included in a Cytogenetics Report by using the KaryoStudio Settings tab You can adjust the number of aberrations included in a Cytogenetics Report by entering a value for MaxReportedD
26. past 4 After you edit the file you must save your changes as a txt file i Illumina recommends saving a copy of the E NOTE original known regions table in the event that you would like to revert back to the default file that is originally provided with the software The Database of Genomic Variants table Figure 28 is a list of all of the CNV regions found in the latest build of the DGV Information from this database is loaded into KaryoStudio as a text file and included for cross matching purposes Found Regions Known Regions DB of Genomic Variants A ol e e sao ll Variation ODU1 Variation D002 Variation 0003 Variation D004 Variation 0005 Variation 0006 Variation 0008 Variation 0009 Variation 0010 Variation 0011 Variation 0012 Variation 0013 Variation 0014 Variation 0015 Variation 0016 na m CTC 232B23 RP1 283E3 RP4 63318 RP11 219C24 RP1 153M3 RP3 423B22 RP6 65F 20 AP11 131015 RP4 501K24 RP11 335D10 RP11 253N12 RP4 580L15 AL021069 RP11 415M14 RP5 1015N21 Amda amma s E de me e AA AA AA AAA AA AA AA 1083805 1609516 8668859 13245159 16880179 37714745 57288465 67485262 83274363 95469800 103956056 113251567 172735904 174297609 231322487 f a mmm ad m 1283805 1720123 8812827 13423743 15338245 37826968 57408242 67607550 83403829 35527423 104113271 113304968 172836399 174457198 231474039 A ton and Figure 28 Database of Genomic Variants 200000 11
27. region In addition if necessary you can edit the start and stop positions of a found region To adjust the confidence score or estimated copy number value of a found region perform the following steps 1 Right click a region of interest in the Found Regions table The context menu appears Figure 53 Part 11328837 Rev B Adjusting Found Region Parameters 71 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 21w122 testC i gliHWisltri2dW OX EB a5x sex 2x sx K 4 N Found Regions i i g Known Regions DB of Genomic Variants Cri du Chat VW Uncheck All B Allele Freq Lider Sampie ID Cry Stat stop length Valu EM UU Bes ee N416593 105587447 105849470 262023 NA16533 64316996 54813520 502524 NA16533 77977306 80668665 2691359 NA16593 7614903 22322881 14707378 NA16593 89665111 83885025 219914 NAI sa 1 47305744 1 47467750 162006 Bay UAOUy EHAJSL ss 288 EE of Genomic Variants Browser Database of Genomic Variants Defects OMIM UCSC Genome Browser PubMed DECIPHER ENSEMBL CHOP Database mm pu aala HHH n o a a tp Io Copy row to Clipboard Copy all to Clipboard 2 00 1 00 0 00 1 00 2 00 Smoothed Log F Copy DBGV CHR Format CO J CO CD Aa a OS Edit a Found Region The project has been loaded successfully Sample Cri du Chat CHR 1 91902273 92863522 Figure 53 Editing a Found Region 2 Select
28. y O wm ES MA Dou uos cO CO CODO C 4 CO n3 B him E fd D A i m LWSdd MdN32 4NG zZ Ndo 4 lxv 297 LOWidd 1 4NG SWAMI 1 Sia 6H0 4 THOS NA16593 91902273 92863522 961249 NA16593 1 147305744 147467750 162006 NA16533 2 89665111 83885025 219914 NA16533 5 7614901 22322881 14707980 NA16593 7 64316996 64819520 502524 NA16593 14 77377306 80668665 2691359 NA16533 14 105587447 105849470 262023 NA16533 16 32349805 32762890 413085 NA16593 x 103148241 103174645 26404 NA16593 x 144917436 144381571 64135 c DA v621S 5 BOB Lv vl 6 5 W403 PET LH axad W495 1 PET LT VI 1483 LINIH 33d zw biNv 4 01404 POPE WN ena 1 had I Baioadaa ss JAM d3dM1 Laide 1 1Sv2 IMJ 1 294 2SUASIL d LONG 1 1 ISSLS Samples ames 1 2 COM HO O t4PJNZ Cri du Chat NA16593 wG0093128 DN Developmental Delay NA 08585 WGO0093128 DNA Wolf Hirschhorn Syndrome N400343 W G0083128 DN Prader Willi rep NA12134 RA W0093128 DN Prader Willi N 12134 WGO0093128 DNA l Mille Dieker Syndrome N 0S208 wG009312 amp DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WGO093128 DNA DO Smoothed Log F Turner Syndrome NA02668 WG0093128 DNA 2dids 1 Yo CEMIS Xi 4 tviseis wr V aro The project has been loaded successfully CHR 5 1 180857866 Figure 49 Found Region Displayed in Context of Whole Chromosome Scrolling Through Found Regions
29. 0 N 15533 7 64316996 64819520 502524 NA16593 14 77977306 80668665 2691359 N 15533 16 32349805 32762890 413085 zx 3 io a gt m iw m o j a zu T Y Mb T E kgs 09 L is 4 P 92 19 SEE P a ue ae L OO E 555 P 92 30 5 giaj C m 1 3 y CT IS S S ST E Ww Samples Cri du Chat NA16593 wG0093128 DN Developmental Delay NA08585 WG0093128 DNA Wolf Hirschhorn Syndrome NA00343 WGO0093128 DNA Prader Willi rep NA12134_A wG0093128 DNA Prader Willi NA12134 WG0093128 DNA Mille Dieker Syndrome NAOS208 wG0093128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WGO0093128 DNA DO Smoothed Log F Turner Syndrome NA02668 WG0093128 DNA TRATE TIT Save is complete Sample Cri du Chat CHR 1 91902273 92863522 Figure 16 Project Created Using a Sample Sheet Continue to Assessing the Quality of Your Data on page 30 tor additional information Loading Perform the following steps if instead of using a sample sheet Sample you prefer to load sample intensity data by selecting the Intensity Data directories that contain the data by Selecting Directories samples in a KaryoStudio project you must create the project using a sample sheet that includes gender data for the samples If you want to include gender data for the NOTE KaryoStudio v1 2 User Guide 22 CHAPTER 2 Loading Your Data 1 From the Choose Sample Loading Method screen click Next to advance to the Loadin
30. 0607 143968 184590 118066 112223 119777 122288 129466 57629 157215 53401 100495 153583 151552 a mM CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber CopyNumber 377 377 8668859 12521755 16612540 37714745 57288465 67485262 83101150 95268437 103762947 112958658 172735904 174297609 230661638 M amma 4d m 2 2 8 1 1 3 5 6 8 3 1 1 1 1 24 b The version of the DGV delivered with KaryoStudio v1 2 is http projects tcag ca variation downloads variation hg18 v8 aug 2009 txt For information about updating KaryoStudio with the latest DGV build see page 87 Part 11328837 Rev B lable 4 Database Column Index ID Landmark Chr Start End Length Variation Type Locus Chr Locus Start Locus End Reference PubMed ID Reference Gain Loss Total Gain Loss Sample Size KaryoStudio v1 2 User Guide Table Window 43 The columns in the Database of Genomic Variants are listed and described in Table 4 of Genomic Variants Columns Description Number used as a simple identifier for regions Identifier assigned to a region by the Database of Genomic Variants stays consistent over time BAC clone identifier for a specific region Chromosome of a region Start position of a region End position of a region Size of a region Type of CNV General ide
31. 1 142400000 153300000 Len 10900000 Detected Region den B Allele Freq CHR 6 00 025 05 075 1 0 Bay puno y O Locus q11 1 Start 61949077 End 62173501 Length 224424 value 3 GL Gain 183 318 2 00 1 00 0 00 1 00 2 00 Smoothed Log R Known Regions 6q pericentromeric region CytoGen65 60500000 63500000 Len 3000000 For Research Use Onky Not for use in diagnostic Dr A J Smith procedures Cornelia de Lange HumanCytoSNP 12v2 1_4 bpm Figure 91 Cytogenetics Report KaryoStudio v1 2 User Guide 114 CHAPTER 6 Generating a Cytogenetics Report Table 9 Cytogenetics Report Header Column Date Sample Name Product Manifest Genome Build cnvPartition Algorithm Version Image of Found Region from Chromosome Browser Description Date the report was generated Name of the sample presented in the report Product manifest used to create the project Genome Build version used for analysis Algorithm version used for analysis not necessarily the most recently installed version Yes Can adjust number of images shown per report Table 10 Cytogenetics Report Detected Region Information Column CHR Chromosome Locus Start End Length Value G L Conf Description Chromosome on which the found region is located Cytoband s on which the found region is located Start coordinate of the found region End coordinate of the found region Length of the found region measured in base
32. 13085 NA16593 103148241 103174645 26404 NA16593 144917436 144381571 64135 Bay uarouy O A 52 puns m M4 B un d gt BG o5 d 3 E uw Ox E s uc B Y Y Z Y o 20819 HH E8451 HHHH LOC WIN QP 11013 He 1038 HHH OO WN wao HH iasa HHH aHa HH cO CO M0 CO P2 C Cri du Chat NA16533 WG0093129 DNA Developmental Delay NAD8585 WG0093128 DNA WolfHirschhom Syndrome NAO0343 WGO093128 DNA Prader Willi_rep NA12134 R WGO0093128 DNA Prader Willi N 12134 wG0093129 DNA Miller Dieker Syndrome NAO09208 WG0093128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WG0093128 DNA DO Smoothed Log F Tumer Syndrome NA02668 W G0093128 DN TOO LD CO J C Cn bon The project has been loaded successfully Sample Cri du Chat CHR 1 91902273 92863522 Figure 24 KaryoStudio Project KaryoStudio v1 2 User Guide 30 CHAPTER 2 Loading Your Data Opening an Existing Project To open a project that was previously created and saved perform the following steps 1 From the KaryoStudio main window click Open an Existing Project 2 Browse to the location of the project you want to open and select Open Your project is loaded into KaryoStudio and the project data displays in the tables and chromosome browser Assessing the Quality of Your Data After your samples are loaded and analyzed in KaryoStudio the software automatically calculates a percent defects score and a LogRDev for each sample These metrics are u
33. 162006 09665111 63605025 213314 105587447 105849470 2523023 32343805 32752830 413085 64316556 64819520 502524 g19502273 92063522 351243 977306 SUBBSKES 2531353 7614301 22322881 14707380 Dil L Figure 50 Aberration Length Sorted Short to Long Click Length again to reverse the sort order The aberrations now display from longest to shortest Figure 51 Part 11328837 Rev B Sorting Found Regions 69 Found Regions Known Regions DE of Genomic Variants E Uncheck All Al Index Sample ID Chr Stat Stop Length Value S ES IS EST S IS T ES ST E Figure 51 ca cO cO Pi om E c c NA15533 NA15533 NA15533 NA15533 NA15533 NA15533 NA15533 NA15533 3 14 1 T 16 14 E 1 NATBSS5 X MA1BS33 X 76143901 POS 305 31302273 54315335 32343805 10558744 09665111 147305744 1443 7435 103148241 2232288 Bl BBSBB55 g2853522 54813521 32762030 105843470 ogogo0z25 147467750 144381571 103174545 14707380 2691 354 361 243 502524 413085 262025 213314 162006 b4135 26404 Aberration Length Sorted Long to Short EXAMPLE 2 Click Chr to sort aberrations based on the number of the chromosome on which they occur Sorting by chromosome is useful if you want to view only aberrations occurring on a certain chromosome Figure 52 Found Regions known Regions DB af Genomic Yarnants WwW Xf Uncheck All Value HA15533 HA1
34. 18 5549719 p subtelomeric region 2 12800000 12800000 Feingold syndrome 2 12500000 17000000 Example Known Regions File Part 11328837 Rev B Appendix D Ignored Regions File Topics 128 Introduction 128 Ignored Regions File KaryoStudio v1 2 User Guide 128 APPENDIX D Ignored Regions File Introduction Table 14 lists and describes the required columns of an Ignored Regions File for use with KaryoStudio Table 14 Ignored Regions File Column Descriptions Column Description Region Name Text description of the region Chr Chromosome number of the region Start Start coordinate of the region Stop Start coordinate of the region The figure below is an example Ignored Regions File Ignored Regions File ad Ignored Regions Table CHR1 5 txE Notepad File Edit Format View Help stop 247249719 242951149 199501827 191273063 18085866 Figure 94 Example Ignored Regions File Part 11328837 Rev B
35. 2 Figure 93 Figure 94 KaryoStudio v1 2 User Guide List of Figures DGV Additional Information 93 DGV PubMed Reference 94 KaryoStudio DGV Table 95 Selecting a Known Region in the DGV Table 95 sienne 97 OMIM HomePage 98 OMIM MYC Gene 99 OMIM Additional Information 100 OMIM Gene Function 101 OMIM Down System Information Page 102 Selecting DECIPHER serle robada gas ad d P Re 105 DECIPHER 554 ws nasa eR REG SR EADEM ES dota eg dou 106 aia o lao aede eb e cem ben adr ee merda d a ear 107 is O 108 Selecting the CHOP CNV Database 109 CHOP CNV Database CNVsS 110 Cytogenetics REDOM era dons de ta 113 Example Sample Sheet 124 Example Known Regions File 126 Example Ignored Regions File 128 xiii Table 1 Table 2 Table 3 Table 4 Table 5 Table 6 Table 7 Table 8 Table 9 Table 10 Table 11 Table 12 Table 13 Table 14 KaryoStudio v1 2 User Guide List of Tables KaryoStudio Toolbar Button Options 35 Found Regions Table Columns 38 Known Regions Table Columns 40 Database of Genomic Variants Colum
36. 2 User Guide 102 CHAPTER 5 Cross Matching to Known Variants Description Clinical Features Cytogenetics Mapping Molecular Genetics t tics R ices Contributors Creation Date Edit History Clinical Synopsis Gene map GenBank Protein U UniGene LinkOut corieLL Figure 84 zx Johns NCBI Hopkins Mv NEU University Sian In Register Genome OMIM OMIM Online Mendelian Inheritance in Man All Databases PubMed Nucleotide swa Sef fer Limits Preview Index History Clipboard Details Display Detailed Show 20 Send to 190685 DOWN SYNDROME GeneTests Links Alternative titles symbols TRISOMY 21 INCLUDED DOWN SYNDROME CHROMOSOME REGION INCLUDED DCR INCLUDED DOWN SYNDROME CRITICAL REGION INCLUDED DSCR INCLUDED TRANSIENT MYELOPROLIFERATIVE DISORDER OF DOWN SYNDROME INCLUDED LEUKEMIA MEGAKARYOBLASTIC OF DOWN SYNDROME INCLUDED Gene map locus Xp11 23 21922 3 1443 TEXT A number sign is used with this entry because the disorder is caused by a chromosomal aberration trisomy 21 and many genes are involved in the phenotype In particular transient myeloproliferative disorder and megakaryoblastic leukemia of Down syndrome are associated with mutations in the GATA1 gene 305371 in conjunction with trisomy 21 y DESCRIPTION Down syndrome the most frequent form of mental retardation caused by a microscopically demonstrable chromosomal aberration is
37. 20636537 Agilent Human 4x44K Patient View gt ec E m e cytoview P ul CO mn e cytoview O co eo e cytoview oO co co e cytoview el cytoview e cytoview e cytoview e cytoview e cytoview Figure 86 DECIPHER Ensembl KaryoStudio allows you to examine your found region within the cytogenetics view of the Ensembl Genome Browser Ensembl is a useful database for examining characterized disorders that may have been previously linked to your region Part 11328837 Rev B Ensembl 107 Using Ensembl To use Ensembl with Karyostudio do the following with Right click a found region and select Ensembl from the KaryoStudio context menu Figure 87 Found Regions Known Regions DB of Genomic Variants des Cont Commen 1520171103 A 1 132721008 192897935 176927 3 78 94658 1520171103 A 7 71728216 73934540 2206324 gt 896 1873 152017711094 8 7403084 31 988585 RETO 6444909 379 8358 485 4874 712 3133 138 1855 245 6331 100 6555 630 609 3456 107 137 3544 117 5533 2611 91 d Database of Genomic variants Browser Database of Genomic variants Defects OMIM LICSC genome browser PubMed DECIPHER Copy row to Clipboard ARAN E E Copy all to Clipboard Copy DEY CHR Format Edit a Found Region Figure 87 Selecting Ensembl Karyostudio automatically sends information about your region to the Ensembl database and the Ensembl Human CytoView win
38. 273 92863522 961249 123 4684 p22 2 p22 1 TGFBR3 BRD NA16593 147305744 147467750 162006 47 05313 q21 1 NA16593 89665111 89885025 219914 48 5083 p11 2 19 NA16593 7614901 22322881 14707380 5318 188 p15 31 p15 2p151p143 1921 ADCY2 NM_0 NA16593 64316996 64819520 502524 247 2509 q11 21 64 ZNF92 NA16593 77977306 80668665 2691359 727 7483 q24 3 q31 1 219 NRXN3 NM N 16533 105587447 105849470 262023 44 71893 q32 33 10 NA16593 32343805 32762890 413085 56 46845 p11 2 19 NM_0010996 NA16593 103148241 103174645 26404 49 39968 q22 2 5 H2BFWT NA16593 144917436 144981571 64135 36 92397 q27 3 12 Mo 2 a2 2 3 2 4 as He 27 He 2 5 oon MN Aa CO n2 2 Figure 26 Found Regions Table KaryoStudio v1 2 User Guide 38 CHAPTER 3 User Interface The Filter Table and Clear Filter buttons allow you to filter the found regions that appear in the Found Regions table and track For more information see Filtering Found Regions on page 63 The Check All Uncheck All buttons allow you to include or exclude all displayed found regions from cytogenetics reports The columns in the Found Regions Table are listed and described in Table 2 Table 2 Found Regions Table Columns Column Index Sample ID Chr Start Stop Length Value Conf Description Identifier given to each region identified by the cnvPartition algorithm across the entire KaryoStudio project all samples Sample name If you use a samp
39. 28678592 133813 8 Conrad et al 200590 A M Figure 69 All CNVs The All CNVs section shows colored bars representing all CNVs present in the database for the genomic region in the view 4 Inspect a variation entry by clicking it A new window opens Figure 70 with details about the selected entry including variation number e cytogenetic band e genes e other overlapping CNVs e genomic coordinates nearby segmental duplications e study details references to the study that reported the variant Part 11328837 Rev B Database of Genomic Variants DGV 91 Database of Genomic Variants Back Home Field Definitions human genome build 36 Variation Variation 3544 Genome context see the graphic below chr5 13300k 13400k Cytogenetic Bands 5pi5 2 All CNVs Blue Loss Red Gain Green GainlLoss Variation_2568 chr5 13197682 13401985 Redon et al 2006 Variation 51574 chr5 1342039 AAA IT Variation 32702 chr5 13204416 13207442 Perru et al 2008 Variation 36 csl Variation 3544 chr5 13218142 13456575 Redon et al 2006 lette 10094 eee doa ZI E AE et al 2007 InDels 188bp to lt 1Kb BluetLoss Red Gain Green GainILoss Variation 11247 chr5 13328913 13329073 de Smith et al 2007 Landmark chr5 13 218 142 13 456 575 Genome Browsers UCSC Ensembl Genomic Position chr5 13 218 142 13 456 575 Variation Type CopyNu
40. 48143238 chrx 48142292 48177927 chrx 488563143 48904371 chrX 48864372 48904850 abe A00NNADID 10044001 N amp 16593 NA08585 e LXXXXXXXXXXXXXX 1G0093128 DNA WG0093128 DNA Cri du Chat B Allele Freq 00 02505 0751 0 15323087 15345343 15422867 15422873 17333868 45433788 46665235 46671151 46695750 46705439 48103430 48142292 48863143 ae AoQnna b Bay puno y Bay UALOUM AO t i J WO pH 9SSOLLLOO WN awa H NA00343 NA12134 R NA 12134 N409208 NA D4250 NAD2658 WG00931 28 DNA WG00931 28 DNA 14G0093128 DNA WG0093128 DNA WG00931 28 DNA WG00931 28 DNA Wolf Hirschhorm Syndrome Prader Willi_rep Prader Willi Miller Dieker Syndrome Retinoblastoma Turner Syndrome 1 A T T T 7 T T 3 T tj n A 3 j 3 y J 3 T J HSNO 44a STNO H send 433144 send Ha E A shee ee 2 00 1 00 0 00 1 00 2 00 Smoothed Log F The project has been loaded successfully Sample Cri du Chat CHR X 153230873 153305225 Figure 78 Selecting a Known Region in the DGV Table You can use this option if you prefer to view CNV data within KaryoStudio rather than linking out to the DGV KaryoStudio v1 2 User Guide 96 CHAPTER 5 Cross Matching to Known Variants Updating the DGV Table and Track The advantage of viewing your data in KaryoStudio is that you can look at a CNV region across multiple samples To update the data used to populate the DGV table and tr
41. 5 219914 NA16593 5 7614901 22322881 14707980 NA16593 7 64316996 64819520 502524 NA16593 14 77977306 80668665 2531353 1 NA16593 14 105587447 105849470 262023 N 16533 16 32349805 32762890 413085 NA16593 x 103148241 103174645 26404 N 15533 x 144917436 144981571 64135 Samples Nam Cri du Chat WGO093128 DNA hi Developmental Delay NA08585 WG0093128 DNA y Weolf Hirschhorn Syndrome N400343 WG0093128 DNA J Prader Willi_rep NA12134_R WG0093128 DNA F Prader Willi N 12134 W G0093128 DNA A Mille Dieker Syndrome NA09208 wG0093128 DNA CO 2 00 1 000 00 1 00 2 00 Retinoblastoma NA04250 WGO0093128 DN4 DO Smoothed Log F Turner Syndrome NA02668 WGO0093128 DNA CO J C CH e w SS The project has been loaded successfully Sample Cri du Chat CHR 5 7614901 22322881 Figure 48 Found Region Displayed in Chromosome Browser If you want to view the selected aberration in relation to the chromosome in which it appears click Expand to Chromosome Intensity data for the whole chromosome is displayed Figure 49 Part 11328837 Rev B Scrolling Through Found Regions 67 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 v122 testC i d id ilc OR OBS EB t 1 5x 1 2x 2x 5x M 4 5 DI Found Regions Known Regions DB of Genomic Variants Y Y Uncheck All B Allele Freq Index Sample lo cm Stat sop leh vaen 02505 07510 1 Bay puno
42. 5533 HA15533 NA15533 NA15533 416535 416533 4165393 N 165353 416533 147300744 31302273 105587447 297306 32343805 63665111 26143901 54315335 103148241 144317435 14467750 32853527 1058434 7l SUBBSEES 32762030 63005025 2232288 54813521 10174645 144381571 162006 361 243 262023 2691359 413065 219914 14707980 5025 24 26404 64135 EE EE 4 5 4 7 4 2 4 3 7 4 8 4 3 Figure 52 Aberrations Sorted by Chromosome When you choose to sort the data for all samples are sorted together Therefore found regions from multiple samples may be intermixed NOTE KaryoStudio v1 2 User Guide 70 CHAPTER 4 Analyzing Found Regions Adding or Editing Comments To add comments or edit existing comments for a found region enter or change a comment in the Comments field For example you can type a note for a colleague suggesting that he or she follows up on this region later You may also want to enter additional information obtained from your cross matching results The comments you enter are included in the Comments section of the pdf reports you generate Adjusting Found Region Parameters Editing Confidence Score or Estimated Copy Number Though it is not generally necessary to do so KaryoStudio offers the ability to edit some parameters of a found region such as confidence score and estimated copy number value KaryoStudio also gives you the option to enter comments for a found
43. 7 Rev B Creating a New Project 7 The Welcome screen appears Figure 4 KaryoStudio Project Wizard Welcome KaryoStudio Project so Welcome to Ehe Karyostudio Project Wizard ul m n a Welcome to Ehe Karyostudio Project Wizard This wizard will quide you through Ehe steps needed to create a project to analyze Illumina Infinium data For structural changes This analysis module supports Illumina Infiniurn Products To load a project you will need a directory containing intensity Files idats a beadpool manifest born and a cluster File egt 4 cluster File is required since all calculations are generated by comparing to normal samples with twa copies Cancel Back Mest gt Finish Figure 4 Wizard Welcome Screen 2 Click Next KaryoStudio v1 2 User Guide 10 CHAPTER 2 Loading Your Data The Project Location screen appears Figure 5 KaryoStudio Project Wizard Project Location KaryoStudio Project 220 www Please specify the name and location for your project U m n a Projects Repository Browse Project Mame m Project will be created in Cancel lt Back Mext gt Finish Figure 5 Wizard Project Location Screen Part 11328837 Rev B Creating a New Project 11 3 To select the repository folder where you would like to store your project do one of the following e Enter a location in the Project Repository field e Browse to a loca
44. 8 CHAPTER 4 Analyzing Found Regions Cri du Chat B Allele Freq 00 02505 0751 0 Bay puno y Rasy UALOUH AO 2 00 1 00 0 00 1 00 2 00 Smoothed Log F Figure 59 Using the Horizontal Ruler 5 Mouse over the ruler at any location to display a tooltip with the location coordinates Figure 60 Cri du Chat B Allele Freq 00 02505 0751 0 Bay puno y Hay LLAAOUM AO ce E C h 7 64 466 623 2 00 1 00 0 00 1 00 2 00 Smoothed Log F Figure 60 Ruler with Coordinates Tooltip Part 11328837 Rev B Adjusting Found Region Parameters 79 In most cases the start and stop positions identified by the cnvPartition algorithm are very close to the positions you can identify by visual inspection However at this resolution you have the option to edit the boundaries of the found region 6 To edit the boundaries of a found region right click the found region of interest in the Found Regions table A context menu appears Figure 61 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_ 1 2 v122 test yg d Wild QR B5 1 sx 1 2x 2x sx H 4 N Found Regions Known Regions DB of Genomic Variants enmarca Y Y Uncheck All B Allele Freq index Sample 0 Chr Stat Step Length Vae Conj 90 02505 07510 6 NA16533 14 105587447 105849470 262023 1 447 4 NAI6593 7 64316996 64819520 502524 1 247 5 N I6533 14 77377306 80668665 2691359 1 727 E
45. A16533 773977306 80668665 2691359 NA16533 7614901 22322881 14707980 N416593 89665111 89885025 213314 NA16533 147305744 147467750 162006 NA16533 31902273 92863522 961249 NA16533 32343805 32762830 413085 NA 16533 103148241 103174645 26404 NA16533 144317436 144381571 54135 Bay puno Hay uA amp OUM O cO CO J O nro Co da mm Samples Cri du Chat NA16593 WG0093128 DNA Developmental Delay NADB8585 W 30083128 DN W ol Hirschhor Syndrome NA00343 WGO00931 28 DNA Prader Will_rep NA12134 R WG0093128 DNA Prader Willi NA12134 wG0093128 DNA Mille Dieker Syndrome NA09208 6G0093128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WG0093128 DNA DO Smoothed Log F Turner Syndrome NA02668 W 30083128 DN e M Siete ee ae igi Ji Sting d og uu BE 01 JNz sHvNd E vsvigs HH sisiWvdov B tozr34 P omi B WzSwl 1 SZESIO WN I SOLA SbLPEL2O1 Dgeesrid asolWw3 orgaan 4 Tawa ZNNSN I Z6S0r vIH I Lwsays TINON dvd z4oav H rSS680L00 WN 4 ZONNLO HHH eos va C9 onum Aa CO P2 The project has been loaded successfully CHR 5 3937903 18645973 Figure 56 Panning Up to Examine the Boundaries of a Found Region Notice that the red track indicating the found region has shifted lower on the screen To expand the view so that you can see additional data adjacent to the end of the found region click Pan Down Figure 57 KaryoStudio v1 2 User Guide 76 CHAPTER 4 An
46. A18318 NA11165 N402668 Found Region NA11165 1 144 967 972 146 453 147 value 3 Figure 37 Karyotype Vie Karyotype Yiew KaryoStudio v1 2 User Guide 56 CHAPTER 3 User Interface Illumina KaryoStudio 1 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and SettingsaleonNDesktopKS v1 2 v122 testC id oc O kK s pund Regions Known Regions DB of Genomic Variants Y W Uncheck all ces Sample ID Ov stat Sen Length valu Con NA02668 2714609 NA02668 65042482 NA02668 83072753 NA 11165 61343077 NA18313 19283777 NA18319 16664861 N418319 15321798 NA18319 21056372 SS GIS e e amples Prader Willi Miller Dieker Syndrome Retinoblastoma Turner Syndrome Cornelia de Lange Smith Magenis Syndrome Down Syndrome Wilms Tumor project has been loaded successfully Figure 39 Founa NA11165 144967972 NA 18313 162580056 58356270 88425251 154582606 65509847 146453147 1485175 62173501 19490411 20358567 15674251 21392612 NA12134 NA09208 NA04250 NA 2668 NA amp 11165 NA18313 NA 4532 NAO 427 162845462 265426 55641 661 1 23382769 224424 206634 3693706 352453 336240 Oy 0 10 MM CO WG0093128 DNA WG0093128 DNA WG00931 28 DNA w G0033128 DN W G0033128 DN W G0033128 DN WG00931 28 DNA WG00931 28 DNA 6811 993 799 1281 193 82 2 75 8 1471 1 44 Cornelia de Lange B Allele Freq 00 02505 0 75 1 0 EJ f amp 1 5x 1 2x 2x S
47. Analysis Ji Illumina recommends analyzing control w NOTE samples along with your experimental samples to ensure that there are minimal processing issues This provides a good point of reference in the event that any issues do arise For additional information about how to OC your data please refer to the KaryoStudio FAOs on the web or contact Illumina Technical Support 32 CHAPTER 2 Loading Your Data Part 11328837 Rev B 34 35 31 45 52 56 57 KaryoStudio v1 2 User Guide Chapter 3 User Interface Topics Introduction Main Window Menus Table Window 37 Found Regions Table 39 Known Regions Table 42 Database of Genomic Variants 44 Samples Table Chromosome Browser Karyotype View Duo and Trio View Log Window 34 CHAPTER 3 User Interface Introduction The KaryoStudio user interface provides tools for loading intensity files browsing found regions comparing found regions to known regions and displaying them graphically Figure 25 shows KaryoStudio s screen configuration Current Genome Build Current Version of cnvPartition Algorithm Found Regions we a Illumina Karyo studio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 v122 testC DUA c4 QR OB 5 Found Regions Kriown Regions DB of Genomic Variants Y Y Uncheck All Index Sample Il LE Lg L Stop Lent vete Con NA16533 j 64316996 NA16593 14 77377306 N 16533 5 7614903
48. Browser PubMed DECIPHER ENSEMBL Samples CHOP Database EE Copy row to Clipboard Cri du Chat zd n Developmental Del Copy all to Clipboard Wolf Hirschhorn S PraderWili_rep Copy DBGY CHR Format Prader Willi T FT Miller Dieker Syndri Edit a Found Region 2 00 1 000 00 1 00 2 00 Retinoblastoma NAO04250 WG0093128 DNA DO Smoothed Log F Turner Syndrome N402668 WGO093128 DN4 DO y b HAN Dai uA Us EH B BIBI tt AILI 1 PLTIPMSIITUCNILEDLENILVUVTLITILUTILPEILLFLUJTILPS 31409 4H zcadwz sHvMaG B ZADOW zsz omi I SFLPEL2O1 1 SHIN I asoiva4 I 4B5S0v vlx Tindoy 4 A Fay puno ELILIELII 3 H3 The project has been loaded successfully Sample Cri du Chat CHR 5 7614901 22322881 Figure 67 Selecting the DGV KaryoStudio sends the chromosome start and stop position information of the found region to the DGV which notifies you of how many matches were found and displays a genome browser view of that region Figure 68 Database of Genomic Variants human genome buxld 35 Showing 14 71 Mbp from chr5 positions 7 614 901 to 22 322 881 D Inatructinns Search using a Sequence name gene mame locus or glhir Fandimark The weldcand character is alowed To center on a locaton click the ruler Liso the ScrolVZoom butions lo change magnibeation and position Examples chr UES AAA CATE patel Oat Hide banner Bookmark this Link to Image High res Image Help FORES E Search Landmark o Regi
49. Delay NA08585 WGO0093128 DNA Wiolf Hirschhorn Syndrome N400343 WGO0931 28 DNA Prader Willi rep NA12134 R WGO0093128 DNA Prader Willi NA12134 WwG0093128 DNA Miller Dieker Syndrome NAOS208 WGOO93128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WGO093128 DNA DO Smoothed Log F Turner Syndrome NA02668 WGO093128 DNA cg Rn Goy puno 4 Il MBit SHvNa B Jaws 1 ZADOW ui I 25680100 WN 4 zani HH ems vs SPlLPElLDO71 d SYLW EH P AA IN zs S cs 0k sha HH moa CO CD Aa w PO The project has been loaded successfully Sample Cri du Chat CHR 5 7614901 22322881 Figure 66 Selecting a Found Region Part 11328837 Rev B Database of Genomic Variants DGV 2 Right click the region and select Database of Genomic Variants Browser from the context menu Figure 67 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon DesktopKS_vw1 21v122 testC y Ui d OQ ck EM 3 1 5x 1 2x zx sx HM 4 N Found Regions i i 9 Known Regions DB of Genomic Variants Cri du Chat Y OW Uncheck All B Allele Freq index Sampleto Chr Stat Stop ergh vane con 09 02505 07510 NA15533 1 31902273 92863522 961249 3 NA16593 1 147305744 147467750 162006 1 NA16593 2 89665111 83885025 219914 1 NA16593 NAT5533 Database of Genomic Variants Browser NA16593 NA16593 Database of Genomic Variants Defects NA16593 OMIM NA16593 NA16593 NA 16533 O UCSC Genome
50. Dieker Sy Copy all to Clipboard 2 00 1 00 0 00 1 00 2 00 Retinoblastoma Copy DBGY CHR Format Smodihed Log F Turner Syndrom Edit a Found Region The project has been loaded successfully Sample Cri du Chat CHR 1 91902273 92863522 Figure 65 Copying Found Regions to the Clipboard You can now paste this data into an Excel file or import it into other downstream third party applications The checkboxes to the left of the Index numbers indicate whether or not a found region will be included in your report All checkboxes are selected by default which means that all found regions will be included in your report by default To deselect all regions click Uncheck All To select all regions click Check All To exclude certain found regions from your report deselect the checkbox es to the left of the region s you want to exclude KaryoStudio v1 2 User Guide 84 CHAPTER 4 Analyzing Found Regions Part 11328837 Rev B 86 87 96 102 104 104 106 108 KaryoStudio v1 2 User Guide Chapter 5 Cross Matching to Known Variants Topics Introduction Database of Genomic Variants DGV OMIM Online Mendelian Inheritance of Man UCSC Genome Browser PubMed DECIPHER Ensembl CHOP Database 86 CHAPTER 5 Cross Matching to Known Variants Introduction Because a sample may potentially contain many aberrations it is especially important to know what has already been identified as a normal variation
51. Edit a Found Region from the context menu The FormSettings screen appears Figure 54 KaryoStudio v1 2 User Guide 72 CHAPTER 4 Analyzing Found Regions FormSettings El Editable Comment Confidence Enabled Start Stop Value El Misc Chr Cytoband CytobandLocus AllteredO ut GainUrLoss Genesh ame Image Length Loci 123 468353 True 91902273 92863522 Gain WA TGFBR3 BRHDT BRADT 5BHD 7 ETB none 961249 Illumina Interface Locus Figure 54 Form Settings Dialog Box 3 Do one of the following e If you would like to change the confidence value for this region adjust the Confidence score E NOTE Although it is not recommend to change the confidence score of a found region you can change it to O if you want to exclude this found region from analysis e Adjust the Estimated Copy Number Value if on visual inspection you feel that cnvPartition has not accurately estimated the copy number of the found region T Y NOTE The parameters in the Misc area cannot be adjusted Part 11328837 Rev B Editing Start and Stop Positions Adjusting Found Region Parameters 73 You can verify the accuracy of the cnvPartition algorithm by examining the start and stop positions of the found region In some cases you may want to adjust start and stop positions ased on visual inspection though it is not common to do so KaryoStudio also allows you to optionally adjust the information in the
52. Found Regions table b CAUTION KaryoStudio v1 2 User Guide If you edit the start and stop positions of a found region the original positions identified by the cnvPartition algorithm are not stored with the project The confidence score number of SNPs and genes do not update after you have edited a region If you need to retain the original start and stop positions for any reason copy the Found Regions table before you load a project and save it as a separate file or save the project on your computer with a different file name CHAPTER 4 Analyzing Found Regions To adjust the start and stop positions of a found region of interest 1 In the Found Regions table select a region of interest Figure 55 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 v122 testC uidi OX E t 1 sx 1 2x 2x sx K 4 gt DW Found Regions i i i g Known Regions DB of Genomic Variants Cri du Chat 57 Y Uncheck All B Allele Freq index Sample ID Chr Stat Step Length Value Con 00 02505 07510 NA16593 105587447 105849470 262023 NA16593 64316996 64819520 502524 NA16593 77377306 80668665 2691359 NA16593 7614901 22322881 14707980 NA16593 89665111 83885025 219914 NA16593 147305744 147467750 162006 NA16593 31302273 92863522 961249 NA16593 32349805 32762890 413085 NA16593 103148241 103174645 26404 NA16593 144917436 144981571 64135 CALS Y tO
53. Gene map locus 3912 1 M 6 164850 Links V MYC AVIAN MYELOCYTOMATOSIS VIRAL ONCOGENE HOMOLOG 1 LUNG CARCINOMA DERIVED MYCLI Homology Maps 3 Gene map locus 1 p34 3 Coriell Figure 81 OMIM MYC Gene If you want to look at this gene in the human genome only for example you must scroll down to find it in the list If you click on a hit information about this gene is displayed including the name description etc KaryoStudio v1 2 User Guide 100 CHAPTER 5 Cross Matching to Known Variants Cloning Gene Function Gene Structure Mapping Edit History Gene map tre En WNome G Protein U UniGene LinkOut Figure 82 OMIM B5 My NCBI a Hopkins Online Mendelian Inheritance in Man y University Sign In Register All Databases Pub led Nucleotide Protein Structure OMIM Search OMIM s for Go Clear Preview Index Clipboard Display Detailed Show 20 y Send to 187270 TELOMERASE REVERSE TRANSCRIPTASE TERT GeneTests Links Alternative titles symbols TELOMERASE CATALYTIC SUBUNIT TCS1 EST2 CORONARY ARTERY DISEASE SUSCEPTIBILITY TO INCLUDED Gene map locus 5p 15 33 TEXT DESCRIPTION Human telomeres consist of many kilobases of TTAGGG n together with various associated proteins Small amounts of these terminal sequences are lost from the tips of the chromosomes each S phase because of incomplete DNA replication but de novo addition of TTAGGG repeats by the enzyme
54. IE KIAAO3O ORIEL p MGC29871 LOC4DD566 SCARFi M PAFAHIBI MM 001114118 NUPB MK RPHSAL TUSCS SRR NM Di4568 fi UBE2G1 W NM D 2 00 1 00 0 00 1 00 2 00 Smoothed Log E Figure 32 Cytoband Tooltip Part 11328837 Rev B Chromosome Browser 49 If you place the cursor over a region found in the DGV track a DGV tooltip is displayed Figure 33 Miller Dieker B Allele Freq Bay pura y Ba UAMOUM AZ o 025 05 073 1 0 9500307 BIOL DO MN Farc dd LANH d Mame Yaristion 4021 Landmark chr17 568336 1008155 Mame variation 32793 Landmark RP11 747K6 Mame Variation 5332 Landmark chr17 60543 963131 Mame variation 8833 gJdMd eras kaos raodo d cJMId3s CvEPOTS M MxN A LTMPMIS H ZZRIL Variation 30020 Landmark chr17 870666 357228 Mame Variation 35522 Landmark chr17 912409 318931 Mame Variation 35521 Landmark chri7 208902 916937 Lobo crio d W ozgan eb be bb LOC car 1 LX I IN OWN sana i t in el E zu m E lo e a e en th e D lo m E zx A vidggAW S6E 00LL00 Wh BH JMH Ewzdlw 4 zalo LEGEZIOIA d es2 pre LLOO PIN aldo 14327 tivo IAH Cds zasgaAs coo Hb 2 00 1 00 0 00 1 00 2 00 Smoothed Log R Figure 33 DGV Tooltip The DGV track is multicolored The colors indicate the numbers of DGV variation regions overlapping at a given point as described below
55. If you want to export data from the Found Regions table you can export a single row of data or the entire table To export a single row Right click in the Found Regions table and select Copy Row to Clipboard To export the whole table Right click in the Found Regions table and select Copy All to Clipboard Figure 65 Part 11328837 Rev B Exporting Found Regions 83 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin Y2 4 4 0 C Documents and Settings aleon DesktopiKS_v1 21v122 testC Qu d c4 OK E ft 1 5x 1 2x 2x 5x M 4 N Found Regions Known Regions DB of Genomic Variants YW Y Uncheck All B Allele Freq Index SamplelD Chr Stat Stop Lengh Walue Con 00 02505 07510 NA16593 14 105587447 105849470 262023 NA16593 7 64316996 54813520 502524 NA16593 77977306 80668665 2691359 NA16593 7614903 22322881 14707378 NA16593 89665111 89885025 219914 NA16593 147305744 147467750 162006 N416593 NA16593 8 NA16593 Database of Genomic Yariants Defects 9 NA16593 OMIM UCSC Genome Browser PubMed DECIPHER ENSEMBL Bay UAOUy O EN F punoJ N d T T T T T AAA SS Be d HM th d d X Bs E q 89 5 8 E o 8 x calo HH sagig WH caso 23148 Loo WN I oi H4 iasa HHH oo WN 3 nao HH 1qsg HHK yo 1 0Ha HH Database of Genomic Yariants Browser Samples Cri du Chat Developmental CHOP Database Wolf Hirschhorm Prader Willi_rep Copy row to Clipboard Prader Willi A RENO RE Miller
56. Long 68 Aberration Length Sorted Long to Short 69 Aberrations Sorted by Chromosome 69 Editing a Found Region 71 Form Settings DialogBok 72 Selecting a Found Region of Interest 74 Panning Up to Examine the Boundaries of a Found Region 75 Panning Down to Examine the Boundaries of a Found Region 76 Centering a Found Region 77 Using the Horizontal Ruler 78 Ruler with Coordinates Tooltip 78 Editing a Found Region 72 Editing the Parameters of a Found Region 80 Form Settings Screen IIIA eee 81 Viewing the New Found Region Positions 82 Copying Found Regions to the Clipboard 83 Selecting a Found Region 88 Selecting the A 89 DGV Genome Browser View 89 LER eones dea ae 90 INSPecinmo a vanao RC KI IIIA 91 DGV Variation Number 91 DGV Cytogenetic Band 92 pne wipe Tem 92 DOV Indels usada soe Roda daw RUP aD dN Ur uN AEE 93 Part 11328837 Rev B Figure 75 Figure 76 Figure 77 Figure 78 Figure 79 Figure 80 Figure 81 Figure 82 Figure 83 Figure 84 Figure 85 Figure 86 Figure 87 Figure 88 Figure 89 Figure 90 Figure 91 Figure 9
57. Once your data has loaded you can view each found region one by one There are multiple ways to select data to display in the chromosome browser Click a found region to display it in the chromosome browser Use the blue arrow buttons IW_ 4 M to move up or down in the Found Regions table gt Use your mouse wheel to scroll up or down the Found Regions table Found regions are only displayed for the sample s selected in the Samples table You can choose multiple samples by Ctrl clicking or Shift clicking multiple rows in the Samples table KaryoStudio v1 2 User Guide 68 CHAPTER 4 Analyzing Found Regions Sorting Found Regions In addition to clicking and scrolling in the Found Regions table you can sort data based on whichever column you like To sort data click a column header NOTE If your found regions table contains hundreds of aberrations it may take some time for KaryoStudio to sort the regions by column header EXAMPLE 1 Click the column header Length which represents the number of bases of an aberration The aberrations listed in this table are now sorted from shortest to longest Figure 50 Found Regions Known Regions DB of Genomic Variants WP Uncheck All sex Senet ch Sat Sep Leng vae Con ca tom c qr cu NA16593 HA15533 HA15533 NHA15533 HA15533 HA15533 HA15533 HA15533 HA15533 HA15533 103148241 264014 14431 7435 64135 147305744 147467750
58. RTS THEREOF OR SOFTWARE OR ANY USE OF SUCH PRODUCT S OUTSIDE THE SCOPE OF THE EXPRESS WRITTEN LICENSES OR PERMISSIONS GRANTED BY ILLUMINA IN CONNECTION WITH CUSTOMER S ACQUISITION OF SUCH PRODUCT S FOR RESEARCH USE ONLY 2010 Illumina Inc All rights reserved Illumina illuminaDx Solexa Making Sense Out of Life Oligator Sentrix GoldenGate GoldenGate Indexing DASL BeadArray Array of Arrays Infinium BeadXpress VeraCode IntelliHyb iSelect CSPro GenomeStudio Genetic Energy and HiSeq are registered trademarks or trademarks of Illumina Inc All other brands and names contained herein are the property of their respective owners Oligonucleotide capture sequences 2003 Illumina Inc All rights reserved Revision History Title Part Number Revision Date KaryoStudio v1 0 Part 11328837 Rev A November 2008 User Guide KaryoStudio v1 2 Part 11328837 Rev B February 2010 User Guide KaryoStudio v1 2 User Guide Chapter 1 Chapter 2 Chapter 3 KaryoStudio v1 2 User Guide Table of Contents A II ea ae Soe li Us o d rr V Table Cr COMICS char ea ok de eder ee peed As vii ISPOPEIOUISB v oru qos sana e que ip qe qe a o dul xi Histor Tables prta aaa es years puto I Pic BOR tna XV OVEIWIOW CD piedad da 1 IIIA PR REESE E Md 2 AudienceandPurpose 2 Installing KaryoStudio 3 KaryoStudio Workflow 6 Loading YourData
59. Table 6 DGV Track Colors Color Number of DGV Variation Regions Overlapping at a Given Point light gold 1 2 orange 3 10 orange red 11 20 red gt 20 KaryoStudio v1 2 User Guide 50 CHAPTER 3 User Interface If you click anywhere along the ideogram an alignment cursor displays as a dashed red line Figure 34 Miller Dieker Syndrome B Allele Freq 00 025 05 O75 1 0 ESSdA HE No AN Y ESSdA MW Lia ros 1 Wind ui 1 ND a Y yO d TveHda W TE Wars 4 dcd3 i ddNI 26140212 CONS AAT rS90vviM d Ll daros 1 GLOW 1 C 3 c7 m uo I Lyi 4 ZWEPOTS bvdd Y 171 Laos zako talo 1 Z LLO 1 cao d yes CON vzdiv d lalHvw3pd h ZYLO 1 l vexo 1 f 7 AD ino MH lav H zas842 1 dee Le OZ Ludo 1432 SAAD 46 tivo EH taani 3an Y 70S i qt LAJANT d oz ESNdS E Owds LMMWv2 ES BA 40212 v IdBaA lave H vd 4 Ydy 4 MON V LT be FP LE H a H H Le H la g H H H H L E ws 2 00 1 00 0 00 1 00 2 00 Smocthed Log R Figure 34 Chromosome Browser Alignment Cursor In the gene display area to the right gene names and transcript locations appear as in the RefSeq database To open RefSeq right click a gene name and select Open RefSeq Part 11328837 Rev B Chromosome Browser 51 The elements of the Chromosome Browser are listed and described
60. Variation 37331 Variation 34392 Variation 3 333 chis 51455333 chis D25340 chis ba 2245 che 52942113 chr 55480842 chis 5548435 chr FUS20842 chis 187639 chis 71 335853 chis 2131414 chis 2152738 chrr z1535353 chr 242456957 24250977 2 51438273 2596558 52067671 23044111 55534500 00006430 0972407 Peor rre 2076104 2182502 2241600 21656391 chrd 403393560 403394560 Figure 77 KaryoStudio DGV Table If you want to see whether any of the found regions in your data match known DGV regions click a region in the DGV table The 51455333 aaa ata 32422461 22342113 559488542 29484351 70820842 7187656392 71336063 2111414 22152730 21595363 24245695 40393560 7 k data for the region you clicked appears in the chromosome browser Figure 78 Found Regions Known Regions DB of Genomic Variants Variation 37355 Variation 37355 Variation 37357 Variation 37358 Variation 37353 Variation 37370 Variation 37371 Variation 37372 Variation 37373 Variation 37374 Variation 37375 Variation 37375 Variation 37377 Variation 37378 it Sahii 27270 Samples Cri du Chat Developmental Delay Landmark chrx 153230873 153305225 chrx 153453493 153475817 chrx 154228672 154416930 chrx 154228791 154417402 chrx 17333868 17339689 chrx 45433788 45475100 chrx 46665235 46709514 chrx 46671151 46702945 chrx 46695750 46756755 chrx 46705439 46744455 chrx 48103430
61. X K 4 b Di a4 m l Ba puno y uaou y A490 2 00 1 00 0 00 1 00 2 00 Smoothed Log F Sample Cornelia de Lange CHR 6 61949077 62173501 Duo and Trio View Found Region Track Region in Chromosome Browser Data plots from multiple samples can be displayed simultaneously in the chromosome browser Duo and trio view can be used to analyze for example a parent parent child trio a control sample or different display settings for two plots from the same sample To use the duo and trio view perform the following steps 1 In the Samples table select the samples you want to display 2 Toadd a second data plot select Settings Trio View Data Plot 2 and select a sample from the dropdown menu Figure 40 Part 11328837 Rev B Duo and Trio View 57 IK 4 b A Data Plot Miller Dieker Syndrome B Allele Frec Genes Display 0 os 0 Data Plot 2 AutoScale LagR Axis Data Plot 3 Settings 417 Figure 40 Selecting Additional Data Plot to Display The second data plot appears in chromosome browser to the right of the original data plot Figure 41 Miller Dieker Syndrome B Allele Freq n n O25 n5 Ds d Miller Dieker Syndrome B Allele Freq n a O25 05 A200 aa a a 23 AJ Ta Eae _ s E E mm 200 4100 O00 100 2 00 200 100 O00 1 00 2 Smoothed Lag E Smoothed Log E Figure 41 Chromosome Browser Duo View 3 Optional To add a third data plot select Sett
62. a use and then selecting directories that contain intensity files From your data repository SNF Manifest O Cree Semper e Browse Data Repository Eres rv Cre Seg Browse Directories in Repository Selected Directories 1520171046 1520171061 1520171061 seer ees 1520171157 1520171109 1520171046 1520171157 Hap3 n Cyto Samples De BeRIDUE Remove All Cancel Back Mext gt Finish Figure 20 Wizard Selected Data 26 CHAPTER 2 Loading Your Data 5 Click Next to advance to the Cluster Positions screen Figure 21 E KaryoStudio Project IF you have an existing cluster File that you want to import i lu m na cluster positions from enter it here Otherwise vou can cluster the samples you ve selected to determine cluster posit T Figure 21 Wizard Cluster Positions Screen Part 11328837 Rev B Creating a New Project 27 6 Browse to the cluster file you want to use with this project Figure 22 KaryoStudio Project Wizard Cluster Positions KaryoStudio Project IF you have an existing cluster file that you want to import Cluster positions From enter it here Otherwise you can cluster the samples you ve selected to determine cluster Figure 22 Wizard Cluster Positions 7 Click Finish KaryoStudio v1 2 User Guide 28 CHAPTER 2 Loading Your Data KaryoStudio displays a progress bar while it loads the intensity files manifest fi
63. able 2 Right click and select OMIM from the context menu Figure 79 Part 11328837 Rev B KaryoStudio v1 2 User Guide OMIM Online Mendelian Inheritance of Man Found Regions Known Regions DB of Genomic Variants Conf E 152017711094 1 152017711094 7 1 za 711 US a 2 1520171061 A 1520171051 4 1520171051 5 1520171045 A 1520171045 1520171045 A 1520171045 A 1520171045 A 1520171045 A 1520171045 A 1520171157 A 132521008 132837335 176927 3 71 28215 73334540 2206324 3 SIEGE oF 403084 d eur Fn 3 78 934558 096 1573 11 56556 EA Ad Database of Genomic Variants Browser Database of Genomic Variants Defects OMIM UCSC genome browser PubMed DECIPHER Ensembl Copy row to Clipboard Copy all to Clipboard Copy DEGY CHR Format Edit a Found Region OMIM opens to the homepage Figure 80 Commen Figure 79 Selecting OMM 97 98 CHAPTER 5 Cross Matching to Known Variants s o Johns 2 NCBI OMIM A ec d Online Mendelian Inheritance in Man eee University Sian In Register All Databases PubMed Nucleotide OMIM Limits Preview Index History Clipboard Details e Enter one or more search terms e Use Limits to restrict your search by search field chromosome and other criteria e Use Index to browse terms found in OMIM records 5 e Use History to retrieve records from previous searches or to combine searches arch Morbid Map OMIM Online Mendelian In
64. ack in KaryoStudio perform the following steps 1 Download the database build from the following location http projects tcag ca variation tableview asp table DGV_Content_Summary txt 2 Download the variation file txt format in the Current Version section based on Build 36 hg 18 of the genome to your computer Example variation file variation hg18 v8 aug 2009 txt Rename this file DatabaseOfGenomicVariation txt Copy the file DatabaseOfGenomicVariation txt replacing the existing file to the directory C Program FilesWlluminal Illumina KaryoStudio 5 Restart KaryoStudio to load the new data file OMIM Online Mendelian Inheritance of Man Using OMIM with KaryoStudio The OMIM database hosted by the National Center for Biotechnology Information NCBI contains information on human genes and genetic phenotypes on all known Mendelian disorders and over 12 000 genes OMIM focuses on the relationship between phenotype and genotype Because OMIM is based upon specific keywords and not basic chromosome start and stop positions KaryoStudio only provides a link to the homepage of this database If KaryoStudio has identified a specific phenotype within the found regions table you can enter a key word e g gene cytoband or phenotype in the homepage of OMIM to obtain more information if available To use OMIM with KaryoStudio perform the following steps 1 Select a region of interest in the Found Regions t
65. also edit this table based on the requirements of your project KaryoStudio v1 2 User Guide 40 CHAPTER 3 User Interface The columns in the Known Regions Table are listed and described in Table 3 Table 3 Known Regions Table Columns Column Index Region Chr Start End Length Disorder Region Description Numeric identifier assigned to known regions Specific name assigned to a known region Chromosome on which the known region appears Position of the first base pair of the known region Position of the last base pair of the known region Length of the known region Name of the dla or the syndrome associated with chromosomal aberrations in this region The default known regions file included m7 NOTE with KaryoStudio is based mainly on affected chromosomal bands so the base pair start and stop positions may be approximate Editing the Known Regions Table Illumina has prepopulated the Known Regions table with a default list of regions of the genome that are of interest to cytogeneticists You can edit or add additional known regions files in order to better represent the regions of interest to your cytogenetics lab You can also have multiple known regions files for different analysis workflows or for use with different sample types Part 11328837 Rev B KaryoStudio v1 2 User Guide Table Window To create a new Known Regions file perform the following steps 1 Create a new file using Microsoft Excel or a
66. alyzing Found Regions Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and SettingsaleonNDesktop KS v1 2v122 testC Dodas E 1 5x sex zx 5x HM 4 OD Found Regi i UEM ound Regions Known Regions DB of Genomic Variants Cri du Chat 7 Y Uncheck All B Allele Freq index SamplelD Chr Stat Stop Length Value Con 90 02505 07510 NA16593 14 105587447 105849470 262023 NA16593 7 64316996 64813520 502524 NA16533 14 77377306 80665665 2691359 NA16533 5 7614901 22322881 14707980 NA16593 2 89665111 83885025 219914 NA16593 1 147305744 147467750 162006 N 16593 1 91902273 92863522 961249 NA16593 16 32349805 32762890 413085 N416593 103148241 103174645 26404 N416593 144917436 144981571 64135 Bay punoy Bay uA OUM ADO A AAA AAA AA A ARA HW HH min MA AO IAN BH II r T 7 X E n o Wd SE E ns Co e a Cn co Co e ra ZLHOO me ldsvg HWNv X4 ZONNLO 81H35 ZZIINZ 4 Sheni cS loer Ol cascoLLOO WN H zslozrid M OLOAW di a 0 00 O PM co Cn amp Samples Cri du Chat NA16583 WwG0083128 DN Developmental Delay NA08585 W G0083128 DN Wolf Hirschhorn Syndrome NAOO343 W G0093128 DN Prader Willi rep NA12134 R WG0093128 DNA Prader Willi NA12134 WGO0093128 DNA l Mille Dieker Syndrome NADI9208 WG0093128 DNA CO 2 00 1 00 0 00 1 00 2 00 Retinoblastoma NA04250 WGO093128 DNA DO Smoothed Log F Tumer Syndrome NA02668 WG0093128 DNA ehasd JO M The
67. ated the cnvPartition algorithm discovers all regions of aberrant copy number based on the cnvPartition configuration settings By default cnvPartition identifies all regions with confidence value greater than 35 and all copy neutral LOH regions larger than 1Mb However these confidence and size thresholds are likely not stringent enough for most uses More specific filtering parameters for size number of markers and confidence threshold can be entered in KaryoStudio After entering the desired parameters only regions of potential interest which meet these criteria are displayed in the Found Regions Table and chromosome browser j For information about adjusting the W NOTE cnvPartition configuration file see the cnvPartition documentation After a project is created filter settings are applied to determine which regions display in the Found Regions Table Default settings are applied to a new project but these parameters can be changed easily To change the filter settings perform the following steps 1 Click 52 Filter Table The FoundRegionsFilterForm appears Figure 47 64 CHAPTER 4 Analyzing Found Regions FoundRegionsFilterForm A ld Use Ignore Regions File Load Known Regione File C Program Files lumina lumina Karnos tudio K Load IhEnownRegionz TypeQfChiy SizeThreshold MarkersT hreshold p CNWConfidenceT hreshold Inside 100000 7 CNLOH 3000000 CNLOH 2000000 Cancel
68. chromosome view to five times closer Adjust chromosome view to two times closer Expand chromosome view by two times Expand chromosome view by five times Jump to the first found region in the Found Regions Table Jump to the previous found region in the Found Regions Table Jump to the next found region in the Found Regions Table Jump to the last found region in the Found Regions Table Adjust KaryoStudio settings for data plots and generating reports Display a log of all actions performed by KaryoStudio Part 11328837 Rev B Table Window 37 Table Window The table window includes three tables Found Regions Table Known Regions Table Database of Genomic Variants These tables are described in the following sections Found Regions The Found Regions table Figure 26 lists the regions passing the Table filter settings that were identified by the cnvParition algorithm found regions and provides the following information Sample name in which a region is found Chromosome on which a region appears Start and stop positions of region Length of region Value Copy Number Conf Confidence Score Comment if any CNV Index Cytobands Number of Markers w v c vV v C VvV VvV vw vV ww Genes Found Regions Known Regions DB of Genomic Variants VW Y Uncheck All under Sample D Chr sien Stop Lent Value Cont Comment CNV Index Cytonands water Genes NA16593 31902
69. d from lllumina s Infinium DNA Analysis BeadChips and generate pdf reports from this data ji KaryoStudio software is for research W NOTE use only Audience and Purpose This guide is written for researchers who want to use lllumina s KaryoStudio software to analyze data generated from IIlumina s Infinium assays to identify cytogenetic or structural aberrations and to generate pdf reports from this data KaryoStudio has been designed specifically for cytogeneticists who are using lllumina s Infinium products to detect aberrations in the genome KaryoStudio accepts Infinium data automatically performs normalization scans data for aberrations displays the analyzed data allowing interpretation by the use and generates reports useful for interpreting results The software is geared towards finding aberrations that are 75kb and larger and cross matching those aberrations with information from public databases Part 11328837 Rev B Installing KaryoStudio 3 KaryoStudio is standalone software application separate from Illumina s GenomeStudio software suite If you would like to obtain comprehensive genotyping information from data generated using Infinium products or perform a more customized data analysis you may want to use the GenomeStudio Genotyping Module For more information about this and other Illumina software products contact your local account manager or visit www illumina com Installing KaryoStudio
70. despread adoption of high density arrays for screening the genome for structural changes many studies have identified various segments of the genome of differing sizes that range in copy number From arrays to sequencing the number of regions is growing dramatically and includes both common gt 5 and rare 1 regions Although many of these regions are not yet linked to specific conditions the hope is that these studies will have a major impact on human health The DGV is one of the databases that is collating these regions in the hope of providing a comprehensive summary of structural variation human genomes This database includes regions of the genome larger than gt 1kb that were identified in various studies of healthy control samples Updating The DGV currently includes many known CNV regions and KaryoStudio covers the majority of the genome with the Latest To update KaryoStudio with the latest DGV build perform the DGV Build following steps 1 Find the latest version of the Genomic Variation Table at the Database of Genomic Variants web site http projects tcag ca variation and click the Downloads section Save the latest version of the Genomic Variation Table as a file of name GenomicVariation txt in the following location C Program FilesWllumina KaryoStudio Restart KaryoStudio to load the updated table Using the DGV You can use KaryoStudio to see if a region identified by with KaryoStudio has already been
71. dow appears Figure 88 KaryoStudio v1 2 User Guide 108 CHAPTER 5 Cross Matching to Known Variants CJ Ensembl Human cyroview e g 4C016942 13 1 152014 AL139815 12 1 95016 Ensembl release 50 HOME BLAST BIOMART SITEMAP HELP ees Your Ensembl El Chromosome 12 Sogn or Register e CHEND 1 15 TO PRO see A ONE E Detailed view Chromosome 12 Features Comparative DAS Sources Repeats Decorations Export Image size 7 852 843 8 017 012 dump to region 12 7es2843 8017012 Bana View of Chromosome 12 Toonicavon EI ELI EIL oo aail anaon A E Graphical overview m Resequencing alignment Length I Forward strand A Kb SN eS Oh 12 bard Paral 726 Mb 790 Mb 792 Mb 794 Mb 796 Mb 798 Mb l NT Contigs NT_009714 4 catarrhini primates 2 Votez Ersembl Genes SLC2A14 UNANOGP1 sic243 rcRNA Genes 23 eutherian mamma 00 omms 0000070 meme 00000 EPO Human tilepath clones as ra Rattus norvegicus DECIPHER No DECIPHER features inthis region Macaca mulatta RedonCNV bei A eee Canis familiaris 786 Mb 728 Mb 790 Mb 792 Mb 794 Mb 796 Mb 798 Mb 8 00 Mb Pan trogiodytes Length 78 M 484 37 Kb Revere strand I MA Mi Ersembi Known Protein Coding MD Meed Known Protein coding m Gallus gallus ae RNA Pseudogene Novel RNA gere Novel Ornithorhynchus There are currertly tracks switched off use the menus above the image to turnthem on anatinus Tupaia belangeri
72. e analysis of all copy number changes in Infinium data B Allele Frequency data is derived trom SNP genotypes while Log R Ratio data is derived from intensity information The B Allele Freq for a sample shows the theta value for a SNP corrected for cluster position Cluster positions are generated from a large set of normal individuals The B Allele Frequency can also be referred to as copy angle or allelic composition It is easier to visualize genotyping data for all SNPs within a chromosomal region using B Allele Freq rather than theta values This is true because B Allele Freq exhibits less locus to locus variation than the theta values for a given sample The transformation of theta values to allele frequencies allows for improved measurements and better visualization of both LOH and copy number changes B allele freq is described by the following equations B allele freq 0 if theta lt tAA 0 5 theta tAA tAB tAA if theta lt tAB 0 5 0 5 theta tAB tBB tAB if theta lt tBB 1 if theta gt tBB where gt tAA mean theta value of all genotypes in the AA cluster 9 yp plotted in polar normalized coordinates tAB mean theta value of all genotypes in the AB cluster plotted in polar normalized coordinates tBB mean theta value of all genotypes in the BB cluster plotted in polar normalized coordinates Part 11328837 Rev B Interpretation KaryoStudio v1 2 User Guide
73. efects On average KaryoStudio includes roughly two aberrations per page so a report with ten aberrations would be approximately five pages long You can limit the number of known regions to display for each found region by entering a value for MaximumNumberOfKnownDefects If you do not want to cross match to any known regions set MaximumNumberOfKnownDefects to 0 Additionally you can select which found regions to include in a Cytogenetics Report by selecting the checkbox to the left of each found region in the Found Regions table Clear the checkbox to the left of a found region to exclude it from a report New Cytogenetics Report After you choose the parameters for the aberration filter and number of aberrations for each report click Write Reports to Directory A dialog box opens allowing you to browse to and select a directory where you want the reports to be saved KaryoStudio generates the reports in the location you selected This takes roughly 30 seconds to one minute per sample Part 11328837 Rev B 118 118 119 120 KaryoStudio v1 2 User Guide Appendix A Additional Information Topics Introduction System Information cnvPartition Algorithm B Allele Frequency and Log R Ratio 118 APPENDIX A Additional Information Introduction This appendix includes information about additional topics that you may be interested in System Information Use the following information as a guide to determine the mi
74. elements of the Log window are listed and described in Table 8 Table 8 Log Window Elements Toolbar Button Element Description if used Select All Selects all log entries E Copy Copies log entries to the clipboard Save Saves all log entries Clear Clears all log entries Es Grid Toggles the grid on and off Time Displays the time the log entry was generated Severity Displays the severity of the log entry Message Displays the text description of the log entry Source Displays the source of the log entry Part 11328837 Rev B 62 63 65 67 68 70 70 82 Chapter 4 Analyzing Found Regions Topics Introduction Filtering Found Regions Displaying Found Regions Scrolling Through Found Regions Sorting Found Regions Adding or Editing Comments Adjusting Found Region Parameters Exporting Found Regions KaryoStudio v1 2 User Guide 62 CHAPTER 4 Analyzing Found Regions Introduction KaryoStudio includes lllumina s cnvPartition algorithm which is designed to scan whole genome Infinium data for intensity changes resulting from changes in copy number Once you have completed entering information into the Project Wizard your project data is automatically scanned using the cnvPartition algorithm Detected aberrations are displayed in the Found Regions table Included in this table is information about each aberration including the sample in which it was found the chromosome start and stop position its size lengt
75. enetics Report 116 Generating a New Cytogenetics Report 116 Additional Information 117 POCO CTO PI 118 System Information 118 cnvPartition Algorithm 119 Conden SCOE cs om os ooh oe AAEE A A EA 119 B Allele Frequency and Log RRatio 120 Calculati n IIIA a ees KAA 120 accio saae s sompa a a aa 2 a AUA 121 Sample Sheet 123 Tae Aa UU edges oie tes ene ce 124 Sample AWA heed 124 Known Regions File 125 ue Po oss IA sue WA AI Tra USE 126 Known Regions File 126 Ignored Regions File 127 INTOCUCION PLC 128 Ignored Regions File 128 ix Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Figure 16 Figure 17 Figure 18 Figure 19 Figure 20 Figure 21 Figure 22 Figure 23 Figure 24 Figure 25 Figure 26 Figure 27 Figure 28 Figure 29 Figure 30 Figure 31 Figure 32 List of Figures Registration Screen 4 Registration Complete 5 Main os ii IIIA IIIA oe 6 Wizard Welcome Screen 9 Wizard Project Location Screen 10 Wizard Project Reposito
76. es display visible select Settings Genes Display Figure 44 Miller Dieker Syndrome B Allele Freq 0 0 025 05 0 75 1 0 O n o a zu T Q Xx a E zu D 2 T de I p V C 3 ay r1 Aq F m m o D 5 a h BE 9LEPLOIOA 1 t0079 LALIAIN SOSL LINI H S1 Lys 6500982501 ITHWA H polo LOO WN Oz bdo 4 13327 LL9GZIOMH E Isi uso C INLMS FTTTWTTTTWTTTFTTTEYTTTTTTTY TT 200 1 00 000 1 00 2 00 LogR Smoothed Log R Figure 44 Single Sample View with Gene Display Log Window The Log window displays information about the projects and tracks for this analysis as they are being loaded into the software Log Of x Time Severi Message 2212010 5 05 13 PM INFO Startup complete 22712070 5 05 18 PM INFO Starting to load C Documents and SettingesaleanD esktopsK S vl Aw 22 testl y1 22 t 2 1 2010 5 05 18 PM INFO Loading manifests 2 1 2010 5 05 22 PM INFO Loading sample Data 4 1 2010 5 05 22 PM INFO Loading sample data 4 1 2010 5 05 22 PM INFO Loading snp data 2 1 2010 5 05 27 PM INFO Copying temp file 2 1 2010 5 05 27 PM INFO Heading Allele Call Index from C Documents and Settingesaleon DresktopsK 5 v1 212 2 1 2010 5 05 27 PM INFO Loading region ao k 4 Last z 5 05 PM Errors ham Info 10 p Figure 45 Log Window KaryoStudio v1 2 User Guide 60 CHAPTER 3 User Interface The
77. frequency for intensity only w NOTE probes is represented by light blue dots and is not used in copy number calculations In addition there is an ideogram of the chromosome found and known regions information from the DGV Database of Genomic Variants and gene information You can adjust the display parameters from the Settings tab To hide or display data types in the plot go to Settings Data Plot and clear or select Smoothed LogR B Allele Freq or LogR The Log R Ratio axes are adjustable By default the axis range is 2 0 to 2 0 To adjust the Log R Ratio axis to fit the data select Settings AutoScale LogR Axis KaryoStudio v1 2 User Guide 46 CHAPTER 3 User Interface Miller Dieker Syndrome B Allele Frey ADO 00 025 05 OFS 10 49d TvcHdH W CCMMIALL 1 MSddMI 2640215 Laos 53d Hi LJMIgs vasi ego 4 dos Ed A LANH LOS NXNM Hal Haa LTJPMIM M Ye HH deus Was dedia tabo tivo EH raans 4 pus zako 1 Lola AO LE pO kingo EH CvEMO Hddzz solo LYEJO LA JMMv Hl LAJMN M Lazaan Y ESMdS i KHa ZSMS l EONI 4 EMMA pasorriH besos 4 ALOLT 2ogds LM ZO ESZLANA ZYEPOTS d Ecza MW Mosia zasg4a2 ijv H zasgro idv H zasao EONI 4 vedi d igipv4vad H dri d T deg AE coc 2m U n ud ex E ERE grs Ludo 1 wd i wd i wd 2 00 1 00 0 00 1 00 2 00 smoothed Lag E Figure 30 Chromosome Browser Search Yo
78. g Sample Intensities screen Figure 17 KaryoStudio Project Wizard Loadin rm KaryoStudio Project Please specify the samples you want to load by identifying the LI ni ci SNP manifest au want to use and then selecting directories i i that contain intensity Files From your data repository SNP Manifest Browse Data Repository Browse Directories in Repository Selected Directories Add gt Remove Remove All Cancel Back Next gt Finish Figure 17 Wizard Loading Sample Intensities Screen Directories Part 11328837 Rev B KaryoStudio v1 2 User Guide Creating a New Project 23 2 Browse to a directory that contains the SNP manifest you want to use with this project Figure 18 KaryoStudio Project Wizard Loading Sample Intensitie KaryoStudio Project Please specify the samples vou wank to load by identifying the U n a SNP manifest you want Ea use and then selecting directories that contain intensity Files From your data repository SNP Manifest trenta rapaz c Sii Browse Data Repository Browse Directories in Repository Selected Directories Remove Remove vB RE Drama al Drama al Cancel VA aa E Back Mext gt Finish Figure 18 Wizard SNP Manifest 24 CHAPTER 2 Loading Your Data 3 Browse to the data repository that contains the data you want to include in this project The direct
79. h in bases an estimate of the copy number Value column a confidence value relative score its cytobands Y v v v v v the number of SNPs it contains the genes present in the found region Once you have created a project all detected regions are displayed in the Found Regions Table and listed in the order in which they were found in each sample Figure 46 Found Regions Known Regions DB of Genomic Variants Y W Uncheck all laten Same DI Chr Stat Sup teng Value Con Comment CNV Indes Cylobands Malen Genes NAI 6533 91902273 32863522 961 249 p22 2 p22 1 TGFBR3 BRADT BROT ABHD BTBD8 Clorfl 45 N 16533 147305744 147467750 162006 q21 1 N 16533 83665111 83885025 219914 p11 2 N 16533 7614901 22322881 14707380 pl5 31 p15 2p15 1p143 1921 ADCY2 NM 001083584 FASTKD3 MTRR SEMA N 16533 64315336 64819520 502524 q11 21 64 ZNF32 NA16593 77977306 80668665 2691359 q24 3 q31 1 219 NRXN3 NM 001105250 DIO2 C1 4orf1 45 TSHR NA16593 105587447 105849470 262023 q32 33 10 NA16593 32343805 32762830 413085 p11 2 19 NM 0010993687 TP53T G 3b NA16593 103148241 103174645 26404 q22 2 5 H2BFWT NA16593 144917436 144981571 64135 q27 3 12 oon MA A CO no c O 0 CD C A CO n2 This chapter describes how to view and analyze the Found Regions in your data Part 11328837 Rev B Filtering Found Regions 63 Filtering Found Regions KaryoStudio v1 2 User Guide When a project is cre
80. heritance in Man Help OMIM Help Jobs ia Welcome to OMIM Online Mendelian Inheritance in Man OMIM is a comprehensive authoritative and timely compendium of human genes and genetic phenotypes The full text referenced overviews in OMIM contain information on all known mendelian disorders and over 12 000 genes OMIM focuses on the relationship between phenotype and genotype It is updated daily and the entries contain copious links to other genetics resources This database was initiated in the early 1960s by Dr Victor A McKusick as a catalog of mendelian traits and disorders entitled Mendelian Inheritance in Man MIM Twelve book editions of MIM were published between 1966 and 1998 The online version OMIM was created in 1985 by a collaboration between the National Library of Medicine and the William H Welch Medical Library at Johns Hopkins It was made generally available on the internet starting in 1987 In 1995 OMIM was developed for the World Wide sE E LIA Web by NCBI the National Center for Biotechnology Information OMIM is authored and edited at the McKusick Nathans Institute of Genetic Medicine Johns Hopkins University School of Medicine under the direction of Dr Ada Hamosh Allied Resources t ce NOTE OMIM is intended for use primarily by physicians and other professionals concerned with genetic disorders by genetics researchers and by advanced students in science and medicine While the OMIM database 1s open to the p
81. iation zhbe s8 chrb 13197682 134015985 Redon et al 20061 Variation 3zy 2 chrB5 1320dd16 13207442 Perru et al 2008 E YVariation_3944 chr95 13218142 13456575 Pedon et al 0067 Variation 10094 chr35 13294912 133858 8 MWang et al C2007 Variation 32703 chr85 13257094 13390719 Perru et al 20057 Variation Soid chr5 13258665 13394742 Pinto et al 2007 Variation 1413 chr5 13267852 133813 8 Lonrad et al 20091 o AZ vn mene H Figure 73 DGV All CNVs In addition start and stop positions of each region and the study in which the region was identified are shown You can click a variation for more information Part 11328837 Rev B Database of Genomic Variants DGV 93 The sections below the CNVs contain information about genomic inversions indels segmental duplications etc In Figure 74 DGV Indels You can click each available section for more information that you may want to incorporate into your cytogenetics study Below the graphic there is some additional information Figure 75 Landmark chra 13 215 142 1345655 Genome Browsers UCSC Ensembl Genomic Position chr5 13 213 142 13456575 Variation Type Copy Number Cytogenetic Band 5p15 2 Starting position along chromosome in Mb 13 2 Gap within 100k No Method BAC Array CGH Reference Redon et al 2006 Pub Med ID 17122550 Frequency Information subject Cohort Control sample Size 270 control samples HapMap Tota
82. igure 62 Editing the Parameters of a Found Region The FormSettings screen gives you the E NOTE flexibility to adjust the Found Regions table a single row at a time Repeat this process for each found region you want to edit 8 Click Start or Stop and enter a new value to the right based on the start or stop position you see in the chromosome browser Part 11328837 Rev B KaryoStudio v1 2 User Guide FormSettings El Editable Comment Confidence Enabled Start Stop Value El Misc Chr Cytoband CytobandLocus AllteredO ut GainUrLoss Genes ame Image Length Lori 9 Click OK Adjusting Found Region Parameters 81 5918 188 True 7614903 22322881 1 5 pl5 31 p15 2 p15 1 p14 3 pl5 31 p14 3 False Loss ADCY NM_OO1089584 FASTKD3 h none 14707380 Illumina Interface Locus Figure 63 Form Settings Screen 10 The results of your change are not immediately visible in the chromosome browser To see the change click a different found region in the Found Regions table Figure 63 82 CHAPTER 4 Analyzing Found Regions Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon DesktopiKS_v1 21v122 testC Quid c4 OQ kas 0D 1 5x 1 2x 2x sx MH 4 b W Found Regions Known Regions DB of Genomic Variants Cri du Chat Y Y Uncheck All B Allele Freq Index Sample ID Chr Start Stop Lengh Value Com 90 02505 07510 Bay pu
83. iller Dieker B Allele Freg 0 75 10 0 25 OS 0 0 mn Son NES ABR MGC14376 TIMM22 a RTN4RL4 IH NXN B SLC43A2 RNMTL1 SERPINF2 Iz un na d a ORIAT M ANKFY1 LUNGS HORIA2 B ANKFYT NM 032 P ORIG CYBSD2 ZFP3 ORID2 K ATP2A3 CHRNE GLOD4 SCARF HE NM 001100398 MYBBP1A GEMN4 PREFS Hg GARNL4 Bp ZZEFI GPIBA FAMS7A P RILP y NM 001098509 CAMKKI H MINKA HI pp13624 SCARF1 4 KI amp ADGE4 ELG NM 001124758 BH NM 001128159 TSRI ORID4 p P2RX1 ARRB FAMIDIB d MYO4C J KIAAD3S7 ORIEL p MGC29671 p LOC400566 SCARFI HY PAFAHTBT MM 001114118 MUPG Ah RPHSAL TUSCS p SRR P NM Di4566 dp UBEZG1 Mp NM D 479 2 593 165 rs106 2 00 1 00 0 00 1 00 2 00 Smoothed Log R SNP Name Tooltip Figure 31 If you place the cursor over a cytoband on the ideogram a tooltip featuring the cytoband name is displayed Figure 32 Miller Dieker B Allele Freq 0 735 1 0 0 25 O 5 0 0 son p ones lp ABR MGC14376 TIMM22 ja RTNARLA IH MXN gp SLO43a2 RNMTL1 SERPIMF2 az rure cna AA PORIAT B ANKFYT UNOS ORIA2 B ANKFYT NM 032 ORIG CYBSD2 4 ZFP3 ORID2 K ATP2A3 CHRNE GLOD4 SCARFI Hg NM_O01100398 MYBBP1A GEMIN4 PRPFS HE GARNL4 gp ZIEF GPIBA FAMS7A p RILP E NM_OO1098509 CAMKKi M MINKI BH pp13524 SCARFi 4 KIAAOBB4 ELG 9 MM DO1124758 BH NM 001128158 TERT p ORID4 P2RX1 ARRE2 FAMIDIB 4 MYC
84. in Table 7 Table 7 Chromosome Browser Elements Element Sample Name B allele frequency plots Log R ratio plot Found Regions Known Regions DGV Chromosome Gene information KaryoStudio v1 2 User Guide Description Shows the sample being displayed Shows B allele frequency data for intensity only markers in blue Shows smoothed log R ratio data in red or log R ratio data in grey Track next to the chromosome showing a found region Gains in copy number are indicated in green and losses are indicated in red Blue boxes represent known regions in this area Directly correlates with the information in the Known Regions Table pto from the Known Regions file currently selected in the ilter Settings table Track next to the chromosome shows correlation with information in the DGV Database of Genomic Variants Vertical ideogram of the chromosome All available RefSeq gene information Note For larger regions such as whole chromosomes the gene information appears very condensed To refine the information use the zoom buttons Pseudoautosomal region PAR markers w NOTE and found regions in the PAR are displayed as being on the X chromosome 52 CHAPTER 3 User Interface Karyotype View The Karyotype View window displays a whole genome view of the found regions for one or more samples This view can be used to gain an initial appreciation for the variation found in a sample or as a clickable visual tab
85. ings Trio View Data Plot 3 and select a sample from the dropdown menu The third data plot appears in chromosome browser to the right of the second data plot Figure 42 KaryoStudio v1 2 User Guide 58 CHAPTER 3 User Interface Miller Dieker B Allele Freq 00 0250 5 07510 Miller Dieker Retinoblastoma B Allele Freq B Allele Freq 00 0250 5 0 75 1 0 00 02505 0 75140 Bay puno y Bay UALDUN 490 2 00 1 000 00 1 00 2 00 m 2 00 1 000 00 1 00 2 00 2 00 1 000 00 1 00 2 00 Smoothed Log Smoothed Log Smoothed Lod Figure 42 Chromosome Browser Trio View To select additional settings for each data plot go to Settings Trio view Data Plot and select or clear the Log R B Allele Freq and or Smoothed Log R checkboxes Figure 43 Miller Dieker Miller Dieker Retinoblastoma B Allele Freq B Allele Freq B Allele Freq Bay puno y ASA AAA Da UOH mm MUDA 00 02505 0 751 0 00 02505 0 751 0 00 02505 0751 0 C h3 c qm e EN C Cn a B i wm B o FTTTUVTTTETTTETTTTTTTETTTTVTTTTVTTTUVTTTS un un gt GO gt 2 z o ho y te G E 2 00 1 000 00 1 00 2 00 2001 000001 002 00 2 00 1 000 00 1 00 2 00 LogR Smoothed Log LogR Smoothed Log LogR Smoothed Log Figure 43 Trio View Additional Plot Settings Selected Part 11328837 Rev B Log Window 59 4 To return to the a single plot view with the gen
86. ings in the FoundRegionsFilterForm click F Save Filter Settings 5 To load settings from a file into the FoundRegionsFilterForm gt Load Filter Settings and browse to the settings file of interest 6 Click OK to apply the new filter settings Displaying Found Regions KaryoStudio v1 2 User Guide Notice that when you select a found region the entire region displays in the chromosome browser Figure 48 By default genotyping information is displayed as B allele frequency blue dots and intensity information is displayed as smoothed log R ratio red line 66 CHAPTER 4 Analyzing Found Regions Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 122 testC QOMGMdA IZZQIKOs 0D t 1 sx 1 2x zx sx HM 4 5 DW Found Regions Known Regions DB of Genomic Variants eric Y Y Uncheck All B Allele Freq index Sample ID Chr Stat stop Lengh Walue Con 90 02505 07510 1 Bay puno y Hay Uanduy ADO Sv4 1 P8S680100 WIN ONNLO HE saul O MN T 6507 IM I TiNdOs 4 JAWS ZADIA 14 SPLRE LOC ES ES S IST S ES ES ST FS E D CO CD C 4 CO nro ce m ii inm c sheng B zstozria 4 omi IK Il laSa HMNv H H TSOYA aso may Hi ANZ ScOLLOO WN S LOZ 4 TC la cca SLHaD HH o NA 15533 91902273 92863522 961249 NA15533 1 147305744 147467750 162006 NA16593 2 89665111 8988502
87. intensity data files After selecting this method continue to Loading Sample Intensity Data by Selecting Directories on page 21 to finish creating the project 14 CHAPTER 2 Loading Your Data Loading Sample Intensity Data Using a Sample Sheet i You must use a sample sheet for optimal NOTE analysis of sek chromosomes KaryoStudio reguires the gender information imported in the sample sheet in order to correctly call found regions or normal regions on the X and Y chromosomes Without a sample sheet data plots will be accurate but found regions may not be Perform the following steps to load sample intensity data using a sample sheet 1 From the Choose Sample Loading Method screen click Next to advance to the Loading Sample Intensities Screen Figure 9 KaryoStudio Project Wizard Loading Sample Intensities KaryoStudio Project aa 22 Please specify the samples you want to load by identifying the U m n a sample sheet and associated data and manifest repositories Sample Sheet Browse Data Repository Browse Manifest Repository Browse Cancel lt Back Next gt Emish Figure 9 Wizard Loading Sample Intensities Screen Sample Sheet Part 11328837 Rev B KaryoStudio v1 2 User Guide Creating a New Project 15 2 Browse to a directory that contains the sample sheet you want to use with this project Figure 18 KaryoStudio Project Wizard
88. is requesting to register the Genome Studio application an thiz Lj Ll m n d machine To register the application please enter your user name and domain below and then select the Register button User Mame Domain aleon LLUMIHA Full Mame Email Alzon Poggi Leon aleancellumina com Company M ame Phone Mumber illurnina Inc g5g 202 4536 Trial Days 1 Figure 2 Registration Complete Click OK KaryoStudio is installed on your computer and opens to the main window Figure 3 5 6 CHAPTER 1 Overview Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin V2 4 4 0 audis 4 O k uu it 1 5x 1 2x 2x sx M 4 gt bl Found Regions Known Regions DB of Genomic Variants WZ v Uncheck All B Allele Freq 00 025 05 075 10 0 1 Sr 3L3t3 JZEdN 4 ESISOLLOO WN 1 PZZSOOLOOD NH Td oz SLOSI acosLO Wh ldvWd 4 ZL1H1M 9123H NANO T6LEON IM 1 PS3H ELA YZLHE v210 XNS gazi ab LOL Samples 290027 1100 WM 4 aWwTvd JF 72 L 17 Er 123 F 48 E Q7 E 0 00 0 00 0 00 0 00 0 00 Smoothed Log R Figure 3 Main Window From this screen you can create a new KaryoStudio project open an existing project view your data generate reports and more Continue to Chapter 2 for information about how to create a KaryoStudio project KaryoStudio Workflow A typical data visualization and analysis workflow using KaryoStudio includes the foll
89. l GainiLoss Related Locus chr5 1319 7652 13466435 This database is implemented using the BIOXRT platform BiaxRT ART viewer version 1 03 Figure 75 DGV Additional Information The frequency information may be particularly useful for interpretation It includes the number of samples studied and the number of times a loss or gain was seen Notice that you can click a link to PubMed if available which displays the publication reference for the study that identified this region Figure 76 KaryoStudio v1 2 User Guide 94 CHAPTER 5 Cross Matching to Known Variants search PubMed E Limia Agvanred search help Publ Ned go od Haberal kiiit el M ji CO 5 Clnar Nahiard lt a Hadr Duspiny Settings Abetrart Sendir E habar Z CH here FAAET Ay Global variation in copy number in the human genome Redon A Ishikawa E Filth KA Few L Perry OHL Andrews TO Fnigber H Shapers MH Carson AR Chen W Cho EE Dallaine 5 Freeman JL Genzaler JA Oralac s M Muang J Kalaitznpnuins D Knmura Cr MacDonald JA Marshall CR Mal A Montgomery L Mishimura Okamura k Shen F Somemile KU Tehinda J Vateecia A Vinnidercark Yang FE Zramg J Zen T Zhang Armengol Conrad DE Este x Tae South C Carter NP Aborted HL Lie Jones KA Seni Siy Fires WA Tha Welcome Trust Larger reitale Welcore Trust Genome Campi Hrodon Combride CEND 154 LE Comenent im Habun AL Mir ih Au 154 tH a Copy number varaton HY ef DHA gequentas i3 fu
90. le and cluster files and runs the cnvPartition algorithm on the data you chose to include in this project Figure 23 karyostudio Progress Stat Loading 1520171046 A Cancel Figure 23 KaryoStudio Status Bar project is directly related to the Infinium product you are using and the number of samples you are processing For example it takes longer to load and analyze data from an Infinium HumanOmni1 Quad BeadChip than it does to load and analyze data from a HumanCytoSNP 12 BeadChip For more information about processing time see the KaryoStudio System Information and Benchmark Performance Technical Note The amount of time it takes to create your NOTE Part 11328837 Rev B Creating a New Project 29 Once the data are finished processing and loading KaryoStudio displays the new project Figure 24 Illumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin V2 4 4 0 C Documents and Settings aleon Desktop KS_v1 2 122 testC ufu OU Et 1 5x 1 2x 2x M 4 gt D ound Regions i i i F gl Known Regions DB of Genomic Variants Cri du Chat 7 Uncheck All B Allele Freq tes Sample DI Chr Stat Stop Length vate Con 00 02505 07510 NA16593 31302273 92863522 961249 NA16593 147305744 147467750 162006 NA16593 89665111 89885025 219914 NA16593 7614901 22322881 14707380 NA 16533 64316996 54813520 502524 NA16533 77977306 80668665 2691359 NA16593 105587447 105849470 262023 NA16593 32349805 32762830 4
91. le frequency sits at O 0 5 and representing the AA AB and BB genotype clusters In regions of the genome that do not have two copies various patterns may be seen You must take into account the direction in which the log R ratio is deflected to determine if a gain or loss of DNA Is present Increases in the log R ratio indicate duplications or amplifications and decreases indicate deletions For more information about interpreting this data see the following Technical Note Interpreting Infinium Assay Data for Whole Genome Structural Variation 122 APPENDIX A Additional Information Part 11328837 Rev B Appendix B Sample Sheet Topics 124 Introduction 124 Sample Sheet KaryoStudio v1 2 User Guide 124 APPENDIX B Sample Sheet Introduction The figure below is an example Sample Sheet for KaryoStudio The following columns are required e Sample ID e SentrikBarcode A e SentrikPosition A gender All other columns are optional If path is not provided in the sample sheet it will be reguired in the new project wizard Sample Sheet Ei Microsoft tecel Sample sheet cry b Ble Edk Mew pet Format onko Data Window Hep Adobe Poe Did ii3 47 aA 412 0 Hd XL fa eos 6 Bon BI UBS s ws Boren rom Eleremen eren spec B dp neg ES window p BRIT arie Al Share As Application HP Weber settings B aj E Header 2 Institute N Genetic Series Investigate Illumina Project Ma HumanCytaSMP 12vt CGentrain
92. le of contents for the found regions in a sample To use the karyotype view perform the following steps 1 In the Samples table right click a sample and select Show Karyotype The Karyotype View window appears Figure 35 All found regions are displayed in a track for each sample selected Regions are color coded e green gain e red loss e purple copy neutral event Part 11328837 Rev B Karyotype View 53 4039200 409200 209208 14039200 409200 409205 I B 08208 409200 4039206 1409205 12039200 409200 509205 409205 409205 409205 409200 409200 409200 NA4039205 4039205 409205 409200 N409200 NADU3208 Figure 35 Karyotype View Window One Sample Selected KaryoStudio v1 2 User Guide CHAPTER 3 User Interface 2 To add or remove additional samples from the Karyotype View press and hold the Ctrl key and click each sample in the Samples table Information for all selected samples appears in the KaryoStudio View window Figure 36 Karyotype View Found Region 299Z0 YN S9LL VN BLES LN 8998CO vN S9 LL VN BIESVVN 8998CO vN S9 LL VN BIES LYN s99Z0T N S9 LL EWN BLES VN S99Z0v N S9 LL VN BLES UTN 999cO v N S9 LL VN BLES LN 899cO v N S9 LL VN BLES LN s99z0v N SOLLL WN BLES LYN 299ZO YN S9LL LN BLES TN 898CO vN S9 LL VVN BIESVVN 9998cO vN S9 LL LvVN BLES LN 999cO vN SOLLLWN BLES LN S99z0v N SOLLLWN BLES LN zzz EEE OD
93. le sheet to load your data sample names are the names assigned to each sample in the sample sheet If you load your data without a sample sheet sample names are barcodes Chromosome number of the found region Position of the first base of a found region Position of the last base of a found region Length in base pairs of a found region Estimate of the physical copy number of a found region O indicates a homozygous deletion loss of both copies 1 indicates a hemizygous deletion loss of one copy e 2 indicates a copy neutral loss of heterozygosity e g UPD or autozygosity 3 indicates a duplication gain of one copy 4 indicates a copy number of 4 or above Confidence score calculated for each found region by the cnvPartition algorithm The confidence score is relative within a iso sample A higher score represents higher confidence alues of 50 or higher tend to reflect regions with high confidence Larger regions tend to have higher confidence scores Part 11328837 Rev B Table Window 39 Table 2 Found Regions Table Columns Column Description Comment User entered comment CNV Index Number assigned to a found region within a specific sample The range begins at 0 Cytobands List of cytobands in the found region Markers Number of markers SNPs or intensity only probes in a found region Genes RefSeq genes present within the found region ji Changing mur selection in the Found NOTE Regions table changes
94. mber Cytogenetic Band 5p15 2 Ctartina nacitian alana zhramacama fim RAY 12 Figure 0 Inspecting a Variation For the purposes of this example Variation_3544 shown above is used Notice the variation number Variation_3544 in the upper left of the screen Figure 71 Database of Genomic Back Home Field Definitions Variation Variation 3544 Genome context see the graphic below Figure 71 DGV Variation Number KaryoStudio v1 2 User Guide 92 CHAPTER 5 Cross Matching to Known Variants This variation number is a permanent number assigned to the region If your lab is interested in this region you can use this ID to refer to this region in the DGV Next notice the cytogenetic band Figure 72 Variation arlation_3544 Genome context see the graphic below chrS 13300k Cytogenetic Bands 5p15 2 EEES All CHVs Blue LozsiRed GainzGreen GainlLozz Variation 25568 chr5 13197582 135401985 Redon et al variation 327 02 chr5 132044156 13207442 Perru et al ii Yvariation_354d4 chr5 13218142 13456575 Redon e Figure 72 DGV Cytogenetic Band This tells you which cytoband your region sits in This is particularly important if you already know that a certain condition is associated with a change in a specific cytoband The next section All CNVs tells you whether any other CNVs have been identified in this region Figure 73 All CHVs i Blue LossiRed GainzGreenzGainlLozz Var
95. ncionat aigniicari but has yel do be f lty ascertainad Wia have canstfructed afirsaenerasam CHY map ofthe human gename mraugh th shidy of 270 indraduals far faur populations with ancestry in Europi rica er Asia m HapMap codec DIA from these individuals was sc need for ORY using bro complementar tec holas SI EA poro prin SHE genotgpang amans aed clone bases comparat ganrt mbrdgesson A Total of IHT copy member variable regiam CAME which exe enrempas overlapping OF adiscer gaina ar lega covering 160 megabases 12 ofthe garicma were identified in these populabons These CHAS contained hundreds of genes disease loci functional elements and segmertal duplications Notably the CANVAS encompassed mann rucleobde content per girai han SMPs understoring fre impose arc NV im pere div amd eeoluben Tho dala eblamed delincalo imps SSequilbrium patos for many CHvg and reves marked variation in copy number among populabone We algo damaonsitrate the utility ofits resource for genet digease studies Pete 17122850 Pubided indexes for MEDLINE 4 Publication Types MeSH Terms Grane Support SEI LinkCha rini rosaancas Figure 76 DGV PubMed Reference The DGV is a resource provided by SickKids Hospital in Toronto mature Rolabod artichon ame ALA op Amr var Miam in i bang or Amero peopled ium Genet 2007 b A genoma ada pursey OF Oop nuni ii Han Chiese needing ir n Tide ZO Cop fumba riribera DONI aT idera ir Ket rra Eriti
96. nimum and recommended system configuration for the computer on which you plan to install KaryoStudio ll N Table 12 System Information NOTE Human 610 Quad 32 bit WinXP SP2 Human CytoSNP 12 Minimum 32 bit System Requirements Operating WinXP SP2 System or Vista Minimum Pentium IV Recommended 1 5 GhZ Processor Pentium IV Requirements 20GhZ NET Version NET 3 5 Network 1 GB Connection Minimum AGB Memory Requirements or Vista Pentium IV 1 5 GhZ Pentium IV 2 0 GhZ INET 3 9 1 GB 4GB The minimum and recommended system configurations vary depending on the products hee plan to use to oad into KaryoStudio data you Human 1M Duo 64 bit WinXP SP2 or Vista Pentium IV 1 5 GhZ Pentium IV 2 0 GhZ NET 3 5 1 GB 8GB enerate the HumanOmnit Quad 64 bit WinXP SP2 or Vista Pentium IV 1 5 GhZ Pentium IV 2 0 GhZ NET 3 5 1 GB 8GB Part 11328837 Rev B cnvPartition Algorithm 119 Table 12 System Information continued Human Human Human HumanOmnit CytoSNP 12 610 Quad 1M Duo Quad Minimum 100GB 100GB 100GB 100GB Hard Drive Requirements Minimum 1024x768 1024x768 1024x768 1024x768 Recommended Video Display 1280x1024 1280x1024 1280x1024 1280x1024 Requirements cnvPartition Algorithm Confidence Score KaryoStudio v1 2 User Guide The cnvPartition algorithm automatically scans your data when you create a new project cnvPartiti
97. no y Bay UALOUN O l 1 I ay canlsv4 1 Jano 1 ZADIA t MIN 1 Sela ovas HH 25680100 WN 1 D14 SYL 21225 lec 10 CO 4 CO ro 6o c o m ii Inm St LEE Loe Fita IS SI ISI S SI FSI ST ST SC lI Oll H TSOYA gsoiWs3 I may Hi ZZ94NZ 4 SHYNO M cswoari ldsva HY H zawwio HH sha HH NA16593 105587447 105849470 262023 NA16593 64316996 64819520 502524 NA16593 77977306 80668665 2691359 NA16593 7614903 22322881 14707978 H NA16593 2 89665111 89885025 219914 NA15533 147305744 147467750 162006 NA18533 31902273 92863522 961249 j NA16593 32349805 32762890 413085 NA16593 103148241 103174645 26404 Re NA16593 144917436 144981571 64135 Samples Name Plate pei Cri du Chat WG0093128 DNA4 i Developmental Delay NA08585 WGO0931 28 DNA y Wolf Hirschhorn Syndrome N400343 WGO00931 28 DNA j Prader Willi_rep NA12134 R wG0083128 DN Prader Willi NA12134 WG0093128 DNA d Mille Dieker Syndrome NAOS208 WGO093128 DNA CO 2 00 1 000 00 1 00 2 00 Retinoblastoma NA04250 WG0093128 DNA DO Smoothed Log F Tumer Syndrome N402668 WGO0093128 DNA Chit Ce Ot tot ee eel AEN J ho The project has been loaded successfully Sample Cri du Chat CHR 5 7614903 22322881 Figure 64 Viewing the New Found Region Positions The new start and stop regions are included in the Found Regions table in the Found Regions track and in all subsequent analyses Exporting Found Regions
98. ns 43 Samples Table Columns 44 DGV Track Colors 49 Chromosome Browser Elements 51 Log Window Elements 60 Cytogenetics Report Header 114 Cytogenetics Report Detected Region Information 114 Cytogenetics Report Footer 115 System Information 118 Known Regions File Column Descriptions 126 Ignored Regions File Column Descriptions 128 Chapter 1 Overview Topics Introduction Audience and Purpose Installing KaryoStudio O Co N N KaryoStudio Workflow KaryoStudio v1 2 User Guide 2 CHAPTER 1 Overview Introduction Cytogenetics is an area of genetics that focuses on chromosomal aberrations resulting in various conditions and phenotypes It includes the routine analysis of G banded chromosomes karyotyping as well as molecular cytogenetics such as fluorescent in situ hybridization FISH and comparative genomic hybridization CGH In recent years many forms of cytogenetics have moved to array based technologies Because of the added benefit that SNP information can provide the field has begun to incorporate such information into their research This user guide describes IIlumina s KaryoStudio cytogenetics software KaryoStudio allows researchers to analyze data generate
99. ntifier that may contain multiple variation IDs Start position of a locus region End position of a locus region Publication reference showing where this region has been published Entrez PubMed ID of a publication linked to a specific region Author of the study Total number of patients in which a gain was seen Total number of patients in which a loss was seen Total number of patients in which a gain or loss was seen Total number of patients in the study u CHAPTER 3 User Interface Samples Table S amples Index Mame 2 3 4 A h T B Figure 29 Samples Tab En du Chat Developmental Delay Wolf Hirschhom Syndrome Prader W ill rep Prader Willi Miller Dieker Syndrome Retinoblastoma Turner Syndrome Comela de Lange Smith h agenis Syndrome Down Syndrome Wilms Tumor The Samples table Figure 29 lists all of the samples you have loaded into KaryoStudio for the current project 416593 46009 ZENA A01 0 65 Female D MA 8585 4460093 22 0N4 AO M ale 18005343 460093 20 0N4 BO Male HA12134 A 46009531205 0H8 B02 Male HA12134 WGO0SS12 DN4 amp CO Male 4039208 460093 20 0N4 CUZ Male 404250 4600931 20 0N4 DO Female MA0zb68 W60093123 0N4 DO Female 311165 44600931 22 0N4 ED Male HA198313 00931 20 0N4 FOI Female 18045392 4400931 20 0N4 GA Female a07427 4600931 20 0N4 H01 Male e When you select a sample in the Samples Table the found regions for that sample appear in the Found Region
100. ome actively regenerating cells such as the stratum basale ofthe skin TERT expression appeared early i during tumorigenesis in vivo beginning with early preinvasive changes in human breast and colon tissues and increasing gradually E D during progression both in the amount of TERT mRNA present within the individual cells and in the number of expressing cells within ith a neoplastic lesion The physiologic expression of TERT within normal epithelial cells that retained proliferative potential and its presence at the earliest stages of tumorigenesis have implications for the regulation of telomerase expression and for the identification of cells that may be targets for malignant transformation y Fossel 1998 reviewed the status of studies examining the relationship between telomerase activity and the aging process as well as the implications of these studies for human health The ectopic expression of telomerase in normal human cells extends their replicative life span Although telomerase expression is a sl 4 44 4 e 4 a4 T n 4 t 4 n 4 4 4 4 4 ESE EN EL R 0 MWST DE mmnm Figure 83 OMIM Gene Function If you enter a key word that is a condition in the OMIM search field OMIM provides information about that condition For example enter down syndrome A listing page comes up with the closest matches Click the first entry 190685 DOWN SYNDROME A down syndrome information page appears Figure 84 KaryoStudio v1
101. on Jens 75143901 22322001 Search Data Source Genome Vananis in Human Genome Build 26 Mar 2005 18 or Scroll Zoom Erw 1471 Mbp El F Flip CDU e eu eeu Cytogenetic Bands CCbrasasaoms meth dolo RefSeq Genes Gene hOCY2 NH 020596 LOCZESEGQ IRE 027117 Diis Hn O0 369 FEL 7 0 2304 LoH39 NH 00334 LDC7Z8411 U mo 4n n Lor fat gH DloUY3S4 CL T5 hn 0320 3 TEIOIW coTI S PEDAL eo bose LUCTZTRITLT E a i ie r Figure 68 DGV Genome Browser View KaryoStudio v1 2 User Guide 89 90 CHAPTER 5 Cross Matching to Known Variants Many optional tracks can be displayed in gt NOTE the DGV browser view Refer to the documentation supplied with the Database of Genomic Variants for more information about using and interpreting information in the browser 3 Scroll down until you get to the section labeled All CNVs Figure 69 All CHVs BluezLossj Red GainiGreen iGainlLoss variation 2b5568 chr5 13197582 134101985 Redon et al 20067 Variation 32 02 chr5 1320d416 13207d42 Perru et al C2000 Variation 39dd chr5 13218142 134565 5 Redon et al 2006 Variation 10094 chr5 13254512 133858 8 Mang et al C2007 Variation 32703 chr35 13257094 13390718 Perru et al 2000 m H A AA Variation 9614 chr5 13258665 13394742 Pinto et al 20075 Variation 1d13 chrs5 13
102. on v2 4 4 provided with KaryoStudio v1 2 has been designed to scan for deletions duplications and amplifications on the autosomes and sex chromosomes The cnvPartition algorithm is based on a recursive partition method cnvPartition which has been optimized for speed and accuracy uses log R intensity and B Allele Freq for identification of chromosomal aberrations estimates copy number values and calculates per locus confidence scores After you load your data into KaryoStudio cnvPartition will automatically scan the data for aberrations For more information about how this algorithm works please see the DNA Copy Number Analysis Algorithms Technical Note at http www illumina com Documents products technotes technote cnv algorithms pdf The confidence score that cnvPartition generates is defined as the sum of all logged likelihoods in the region for the assigned copy number minus the sum of all logged likelihoods of copy number equal to 2 for loci in the region Thus the confidence scores provide a means to rank regions relative to their dis similarity to normal copy number 2 segments Higher values represent higher confidence in the aberration call 120 APPENDIX A Additional Information For more information about recommended confidence score cutoff values see the KaryoStudio FAOs on htto www illumina com B Allele Frequency and Log R Ratio Calculation B Allele Frequency and Log R Ratio data form the basis of th
103. onservation of this gene in mammalian species is shown on a graph Next the conservation of this gene in other species is shown on tracks Finally the location of SNPs in humans and the location of repeated DNA sequences Below all of this you can select various additional information to include in this graph The Phenotype and Disease Association Studies track might be of interest Select these tracks to see if any known phenotypes have been associated with this specific region 104 CHAPTER 5 Cross Matching to Known Variants PubMed Using PubMed with KaryoStudio DECIPHER PubMed available via the NCBI Entrez retrieval system is hosted by the National Center for Biotechnology Information NCBI at the National Library of Medicine NLM at the U S National Institutes of Health NIH PubMed provides access to citations from biomedical literature and is a streamlined way to determine if there are any peer reviewed and published studies related to the region condition or phenotype under study Since this database is based on specific key words not on chromosome number and position KaryoStudio provides only a link to the homepage of this database If KaryoStudio has identified a specific phenotype within the found regions table you can enter the keyword in the homepage of PubMed to obtain more information This section is similar to OMIM in that it is based on keywords Type in a gene name condition name etc
104. ories available for selection appear in the Directories in Repository area Figure 19 KaryoStudio accepts manifests only from a 7 NOTE single Infinium product type You cannot create a KaryoStudio project with multiple products or with different versions of the same product KaryoStudio Project Wizard Loading Sampl Intensit KarvoStudio Project Please specify the samples vou want to load by identifying Ehe Ll n ci SNP manifest you want Ea use and then selecting directories that contain intensity Files From your data repository SNP Manifest O 00 re Sept Browse Data Repository E TestDatalRepositorylKaryoStudiolHap300_Cyto_ Samples Browse zl Directories in Repository Selected Directories 1520171046 1520171061 1520171109 1520171157 Hap300_Cyto_ Samples Demo Deene Remove Remove Ail Cancel trees a aca aa lt Back Next gt Finish Figure 19 Wizard Data Repository Part 11328837 Rev B KaryoStudio v1 2 User Guide Creating a New Project 25 4 Add data to this project by selecting items in the Directories in Repository area and clicking Add The directories you selected appear in the Selected Directories area Figure 20 KaryoStudio Project Wizard Loading Sample Intensibie KaryoStudio Project Please specify the samples vou want to load by identifying the U n a SNP manifest you want E
105. owing steps 1 Use the project wizard to load data into KaryoStudio 2 Scan the data for aberrations using KaryoStudio 3 Adjust the filter settings to display found regions of interest 4 Display found regions of interest in the chromosome browser and Found Regions table 5 Cross match found regions to areas of the genome known to be associated with various conditions or phenotypes 6 Select report settings 7 Generate a KaryoStudio Report Part 11328837 Rev B 30 Chapter 2 Loading Your Data Topics Introduction Creating a New Project 14 Loading Sample Intensity Data Using a Sample Sheet 21 Loading Sample Intensity Data by Selecting Directories Opening an Existing Project 30 Assessing the Quality of Your Data KaryoStudio v1 2 User Guide 8 CHAPTER 2 Loading Your Data Introduction Using the Project Wizard is an easy way to begin a new KaryoStudio project Before you create a project make sure that you have the following files easily accessible in a known location on your computer Intensity data files idat files from a BeadChip or BeadChips of interest A product manifest bpm file specific to the BeadChip product used A product cluster file egt file specific to the BeadChip product used Creating a New Project To create a new KaryoStudio project perform the following steps 1 On the KaryoStudio main screen click New Project to open the Project Wizard Part 1132883
106. pboard Copy all to Clipboard Copy DEY CHR Format Edit a Found Region Prader Willi 4612134 600937 28 06 Miller Dieker Syndrome 409208 600937 20 06 Retinoblastoma 604250 600937 28 06 Turner Syndrome MAUZBBS 14600931 z8 D HA cO 07 CH Es La pa al Figure 89 Selecting the CHOP CNV Database KaryoStudio automatically sends information about the found region to the CHOP Database and opens the CHOP CNV browser window to display any CNVs in the CHOP Database that map to the region you submitted Figure 90 KaryoStudio v1 2 User Guide 110 CHAPTER 5 Cross Matching to Known Variants a Sed settee Ll AS CHY tracks 9 CHOP CNV Blocks CHOP CNVRs R CHOP CVs Toronto CHV s Figure 90 CHOP CNV Database CNVs Part 11328837 Rev B 112 112 116 116 Chapter 6 Generating a Cytogenetics Report Topics Introduction What s in a Cytogenetics Report Adjusting the Information in a Cytogenetics Report Generating a New Cytogenetics Report KaryoStudio v1 2 User Guide 112 CHAPTER 6 Generating a Cytogenetics Report Introduction KaryoStudio allows you to create a pdf Cytogenetics Report for each sample summarizing the aberrations found in that sample and showing whether they cross match with any known regions in the genome Depending upon how many aberrations are found for a sample and how many you choose to include in the report it can be from one page to many pages long You need
107. project has been loaded successfully CHR 5 11291957 26000187 Figure 57 Panning Down to Examine the Boundaries of a Found Region Notice that the red track indicating the found region has shifted higher on the screen ii It is best to pan up until the start of the Ww NOTE found region appears in the center of the chromosome browser or pan down until the end of the found region appears in the center of the chromosome browser so that when you zoom in further the SNPs of interest are visible in the window Once your found region is centered in the chromosome browser you can zoom in to get a higher resolution view of the boundaries of your found region Alternatively you can pan and zoom by selecting and dragging the red box which denotes the found region as described in Navigating the Chromosome Browser on page 47 3 Click Zoom in 2x or Zoom in 5x for a higher resolution view of your found region Figure 58 Part 11328837 Rev B Adjusting Found Region Parameters 77 E If you want to view the boundaries of your yu NOTE found region at the SNP level to see each individual data point in the plot you may need to zoom in multiple times Ilumina KaryoStudio 1 2 2 Build 36 1 CNY Plugin V2 4 4 0 C Documents and Settings aleon DesktopKS_vy1 21wv122 testC iQ cO GEB DA 1 5x 12x 2x SX M 4 gt N Found Regions i i i g Known Regions DE of Genomic Variants Cri du Chat Y Y Uncheck All B Allele Fre
108. q Index Sample 1D Chr Stat Stop Lengh Value Conj 90 02505 07510 NA16593 105587447 105849470 262023 NA16593 64316996 64819520 502524 NA16593 773977306 80668665 2691359 NA16593 7614901 22322881 14707980 N416593 83665111 89885025 219914 N416593 147305744 147467750 162006 N 15533 31302273 32863522 961249 NA16533 32343805 32762830 413085 NA 16533 103148241 103174645 26404 NA16593 144917436 144981571 64135 Bay puno y C ESTA j2 1S El WN vH EC Vv vM 6 BSONN 1 poa 131 v VYsdys pe P25680L00 WN 1 Nod PSPs 8 okaga 2zi2d022 aa I ais ZN SGCESLO WN DIEEE Svd QNNIO WF EQMI c 0 CO JO PM 0d Oc OM N SOM 1 gasoil 4 OLOAW 1 sHvNa I c Hand d 2952054400 WN Loza 4 ZZ94NZ Samples SLHaD 4 cS ZLHOO H4 eld 4 Cri du Chat NA15583 wG0093128 DNA Developmental Delay NA08585 WG0093128 DNA Wolf Hirschhorn Syndrome NA00343 W G0083128 DN4 Prader Willi rep NA12134_R WG0093128 DNA Prader Willi NA12134 WG0093128 DNA Mille Dieker Syndrome N 09208 WGOO93128 DNA CO 2 000 1 000 00 1 00 2 00 Retinoblastoma NA04250 G0093128 DNA DO Smoothed Log F Tumer Syndrome NA02668 W G0083128 DNA qo 4 The project has been loaded successfully CHR 5 260730 29677130 Figure 58 Centering a Found Region 4 To display a horizontal ruler click anywhere on the ideogram The horizontal ruler appears Figure 59 KaryoStudio v1 2 User Guide 7
109. ry Location 11 Wizard Project Nami iii 9 REX EIA 12 Wizard Loading Sample Intensities Screen 13 Wizard Loading Sample Intensities Screen Sample Sheet 14 Wizard Sample Sheet 15 Wizard Data Repository 16 Wizard Manifest Repository 17 Wizard Cluster Positions 18 Wizard Select Cluster Positions File 19 Kafryobtudio Status Bal s usce ce tea de et none ds 20 Project Created Using a Sample Sheet 21 Wizard Loading Sample Intensities Screen Directories 22 Wizard SNP Manifest 23 Wizard Data Repository 24 Wizard Selected Data 25 Wizard Cluster Positions Screen 26 Wizard Cluster Positions 27 KaryoStudio Status Bal s aca aor depu cee bear none ds 28 KaryoStudio Project 29 KaryoStudio Screen Configuration 34 Found Regions Table 37 Known REGIONS Table 39 Database of Genomic Variants 42 Samples Table 44 Chromosome Browser 46 SNP Name Tooltip 48
110. s table above You can choose more than one sample by pressing and holding the Ctrl button on your keyboard while selecting multiple samples of interest As you select each sample the Found Regions table above is dynamically updated E NOTE The columns in the Samples Table are listed and described in Table 5 Table 5 Samples Table Columns Column Index Name ID Plate Well Description Number assigned to the sample for sorting purposes Name or barcode of a sample from a BeadChip Barcode of a BeadChip Location on the sample preparation plate Well on the sample preparation plate Part 11328837 Rev B Chromosome Browser 45 Table 5 Samples Table Columns Column Defect Gender LogRDev Chromosome Description Score given to each sample based on the number of found regions This value is the sum of the length of all found regions per sample divided by the length of the genome Gender of the sample as provided in the sample sheet KaryoStudio uses the gender information to correctly interpret found regions on the sex chromosomes Standard deviation of the log R ratios of the sample Browser The Chromosome Browser includes representations of your data in the form of B allele frequency genotyping information represented by blue dots and log R ratio intensity information represented by a grey line The smoothed log R ratio is represented by a red line Figure 30 T The B allele
111. seful when determining whether there are any issues with a specific sample The percent defect score and LogRDev are displayed in the lower left in the Samples Table The percent defects score is calculated by summing the length of all of the found regions within a particular sample of interest and dividing it by the length of the genome In control samples such as non constitutional samples a percent defect score of less than 1 is expected In most cases the value is likely to be less than 0 5 In samples that contain many aberrations such as tumor samples the percent defect score is expected to be much higher in the 60 70 range In addition if there were any issues during the processing of your Infinium products such as a misregistration or sample processing error this score is expected to be abnormally high Part 11328837 Rev B KaryoStudio v1 2 User Guide Assessing the Quality of Your Data 31 The LogRDev metric is a measure of the noise in the data calculated as the standard deviation of the log R ratios for the SNPs on the autosomes This metric is a simple indicator of the variation in the intensities measured for all markers from a particular sample When analyzing relatively normal samples and following standard Illumina protocols for Infinium HD products LogRDev is expected to be less than 0 3 For more information about interpreting LogRDev see the Technical Note Evaluating Infinium Array Data for Copy Number
112. ster the Genome Studio application on this LJ m n a machine To register the application please enter your user name and domain below and then select the Register button User Name Domain Jaleon ILLUMINA Full Marne Email NENNEN Company Mame Phone Number A AAN Trial Days 1 Register Cancel E Figure 1 Registration Screen If you have previously installed and registered another version of KaryoStudio the registration screen does not appear W The Registration Screen has a NOTE GenomeStudio GS icon and refers to Illumina s GenomeStudio software suite This is because KaryoStudio while separate from GenomeStudio uses the same registration database 3 Complete the following information e User Name Enter the user name you use to sign into your computer Part 11328837 Rev B KaryoStudio v1 2 User Guide Installing KaryoStudio e Domain KaryoStudio recognizes your network domain and populates this field automatically e Full Name Enter your full name Email Enter your email address Company Name Enter the name of your organization company institute university etc e Phone Number Enter your phone number ji Illumina uses this information for Technical NOTE Support purposes Click OK KaryoStudio is registered Figure 2 and you can continue the installation process E Registration VM To better serve customers Illumina Inc
113. telomerase compensates for this loss Many human cells progressively lose terminal sequence with cell division a loss that correlates with the apparent absence of telomerase in these cells Kipling 1995 Y CLONING Morin 1989 identified the nbonucleoprotein telomerase in HeLa cells Catalytic subunits of telomerase from the ciliate Euplotes aediculatus and the yeast Saccharomyces cerevisiae contain reverse transcriptase motifs Nakamura et al 1997 identified homologous genes from the fission yeast Schizosaccharomyces pombe and x OMIM Additional Information Of particular interest is the Gene Function section which describes any known molecular function of this gene Figure 83 Part 11328837 Rev B OMIM Online Mendelian Inheritance of Man 101 x Aj GENE FUNCTION Morin 1989 found that human telomerase recognized a single stranded G rich telomere primer and added multiple telomeric repeats to its 3 prime end by using an RNA template in vitro Wilkie et al 1990 found that a case of human alpha thalassemia was caused by a truncation of chromosome 16 that had been healed by the addition of telomeric repeats TTAGGG n Using an in vitro assay Morin 1991 showed that human telomerase correctly recognized the chromosome 16 breakpoint sequence and added the repeats indicated They suggested that telomerase based chromosome healing may stabilize a broken chromosome and allow its stable inheritance The Miller Dieker syndrome
114. tion Figure 6 i 2 KaryoStudio Project LI Please specify the name and location For your project na LEITET Figure 6 Wizard Project Repository Location KaryoStudio v1 2 User Guide 12 CHAPTER 2 Loading Your Data 4 In the Project Name area enter a name for your project in the text field Figure 7 KaryoStudio Project 22 TL Please specify the name and location For your project lu m i na aaa 3 4 Extensa Ali Figure 7 Wizard Project Name Part 11328837 Rev B Creating a New Project 13 5 Click Next to advance to the Loading Sample Intensities screen Figure 8 KaryoStudio Project Please specify the samples you want to load by identifying the LII n a sample sheet and associated data and manifest repositories C Use sample sheet to load sample intensities Load sample intensities by selecting directories with intensity Files Cancel lt Back Mext Eine Figure 8 Wizard Loading Sample Intensities Screen 6 Select one of the following options KaryoStudio v1 2 User Guide Select Use sample sheet to load intensity data if you have a sample sheet that you want to use with this project After selecting this method continue to Loading Sample Intensity Data Using a Sample Sheet on page 14 to finish creating the project Select Load sample intensities by selecting directories with intensity data if you want to load project data directly from
115. u can navigate to a particular location in the chromosome Function browser by entering a search term in the search field Searches may be of the following types gt Chromosomal coordinate Enter the chromosome number a colon and the start and stop coordinates separated by a hyphen EXAMPLE enter 1 1 2000000 to display the first 2Mb of chromosome 1 Part 11328837 Rev B Chromosome Browser 47 gt Cytoband Enter the location in ISCN notation EXAMPLE enter 1p22 1 or 1p22 Gene name Entering text into the search box displays the first gene that matches the search term starting with the chromosome currently displayed in the browser To search a particular chromosome start your search with the chromosome number and a colon EXAMPLE enter 6 VEGF to search for VEGFA rather than VEGFC on chromosome 4 Navigating the In addition to using the toolbar navigation buttons you can Chromosome zoom in and scroll by clicking and dragging the red box on the Browser ideogram using your mouse To scroll click and drag the red box up or down along the ideogram To zoom in our out drag the top or bottom edge of the square up or down along the ideogram The size of the box indicates the size of the chromosomal region displayed in the data plot If you place the cursor over a data point a tooltip featuring the SNP name and coordinates is displayed Figure 31 KaryoStudio v1 2 User Guide CHAPTER 3 48 User Interface M
116. ublic users seeking information about a personal medical or genetic condition are urged to consult with a qualified physician for diagnosis and for answers to personal questions Figure 80 OMIM Home Page 3 Type a key word in the search text field Key words can be gene names etc For example if you had identified the MYC gene in your search you might want to enter MYC as a key word OMIM identifies your gene in multiple organisms and displays a list of the occurrences of this gene in all organisms Figure 81 Part 11328837 Rev B OMIM Online Mendelian Inheritance of Man 99 3 Johns OMIM e UE Online Mendelian Inheritance in Man T University Sign In Register All Databases PubMed Nucleotide Protein Genome Structure OMIM Search OMIM for v vc Go Clear Save Search Preview Index Clipboard Details Eres Display Titles y show 20 y Sendto y Ali 303 OMIM UniSTS 25 OMIM dbSNP 26 s Items 1 20 of 303 Page of 16 Next 1 190080 Mns II V MYC AVIAN MYELOCYTOMATOSIS VIRAL ONCOGENE HOMOLOG MYC Gene map locus 2024 12 q24 13 OMIM Help How to Link Tl 2 164840 GeneTests Links V MYC AVIAN MYELOCYTOMATOSIS VIRAL RELATED ONCOGENE NEUROBLASTOMA DERIVED MYCN Gene map locus 2p24 1 3 606535 MYC BINDING PROTEIN MYCBP Gene map locus 1p33 p32 2 M 4 113970 BURKITT LYMPHOMA BL Gene map locus 3q24 12 q24 13 5 612049 Links MYC INDUCED NUCLEAR ANTIGEN MINA
117. um gt Figure 14 Wizard Select Cluster Positions File 20 CHAPTER 2 Loading Your Data Click Finish KaryoStudio displays a progress bar while it loads the intensity files manifest file and cluster files and runs the cnvPartition algorithm on the data you chose to include in this project Figure 23 amp h KaryoStudio Progress Stal VEN Loading 1520171045 A Figure 15 KaryoStudio Status Bar a The amount of time it takes to create your Ww NOTE project is directly related to the Infinium product you are using and the number of samples you are processing For example it takes longer to load and analyze data from an Infinium HumanOmni1 Quad BeadChip than it does to load and analyze data from a HumanCytoSNP 12 BeadChip For more information about processing time see the KaryoStudio System Information and Benchmark Performance Technical Note Part 11328837 Rev B Creating a New Project 21 Once the data are finished processing and loading KaryoStudio displays the new project Figure 24 Illumina KaryoStudio 1 2 Build 36 1 CNY Plugin 2 4 4 0 C Documents and Settings aleon Desktop KS_vy1 2Project_Info P1 uiid Ok Bi DA sx 172x 2x 5x M 4 N Found Regions i i i 9 Known Regions DB of Genomic Variants Cri du chat Y Y Uncheck All B Allele Freq index Semple Chr Stet Stop lengh Vae Co 00 02505 07510 NA16593 1 91902273 92863522 961249 N 15533 5 7614901 22322881 1470798
118. what is displayed in the Chromosome Browser to the right of the table Known The Known Regions table Figure 27 lists all of the known Regions Table regions that have been loaded into KaryoStudio Found Regions Known Regions DB of Genomic Variants EE AA A SSS ae CytoGeni 27800000 27800000 1p35 Microdeletion CytoGen2 6100000 20300000 4200000 Bartter3 classic CytoGen3 16100000 56200000 40100000 Bartter4 infantile with sensorineural deafness CytoGen4 60900000 68700000 7800000 NFIA Haploinsufficiency CytoGen5 117600000 120700000 3100000 1p pericentromeric region CytoGen6 142400000 148000000 5600000 1921 1 Microdeletion with susceptibility for thrombocytopenia absent radius TAR CytoGen 142400000 153300000 10900000 19 peticentromeric region CytoGen8 174300000 178600000 4300000 Short stature pituitary and cerebellar defects amp small sella turcica CytoGen9 205300000 209500000 4200000 Van der Woude CytoGen10 212100000 222100000 10000000 1941 342 Microdeletion Fryns CytoGen11 234600000 247249719 12649719 1q subtelomeric region CytoGen12 241700000 247249719 5549719 1944 CytoGen13 0 12800000 12800000 2p subtelomeric region CytoGen14 12800000 17000000 4200000 Feingold syndrome CytoGen15 38400000 41600000 3200000 Noonan4 1 2 3 4 5 6 8 3 IN N N me st ll ll Figure 27 Known Regions Table In this table Illumina provides information about commonly affected regions by default However you can
119. with this project Figure 12 KaryoStudio Project Wizard Loading Sample Intensities KaryoStudio Project EEE de Please specify the samples you want to load by identifying Ehe U n a sample sheet and associated data and manifest repositories 60 l Sample Sheet Fc and Settings aleoniDesktopiKESs_v1 2 Project_ Infor Hu Browse Data Repository roce and Settings aleoniDesktopiKES_v1 2 Project_Infor ID Browse Manifest Repository C Documents and Settings aleoniDesktopieS v1 2 Project_Info Cancel lt Back Next gt Finish Figure 12 Wizard Manifest Repository 18 CHAPTER 2 Loading Your Data 5 Click Next to advance to the Cluster Positions screen Figure 13 KaryoStudio Project IF vou have an existing cluster File that you want to import cluster positions From enter it here Otherwise vou can cluster the samples you ve selected to determine cluster posit arien Figure 13 Wizard Cluster Positions Part 11328837 Rev B KaryoStudio v1 2 User Guide Creating a New Project 19 6 Browse to the cluster file you want to use with this project Figure 14 ao a KaryoStudio Project a eo E IF vou have an existing cluster File that you want to import lu na cluster positions From enter it here Otherwise you can cluster the samples you ve selected to determine cluster posit Poona and Settings aleon Desktop KS_v1 2 Project_InfolH
120. word processing program or start with an existing Known Regions file 2 Verify that the required column headers are included see the bulleted list below Enter information in each row for each known region Save the file as a tab delimited file txt in the KaryoStudio program directory The default KaryoStudio program directory is C Program FilesWlluminaKaryoStudio To edit a Known Regions file perform the following steps 1 Navigate to the KaryoStudio program directory The default KaryoStudio program directory is C Program Files IIlumina KaryoStudio 2 Open the known regions file to be edited The default file is KnownRegionsTable txt in a text editing tool such as Microsoft Excel Any of the rows in this file can be edited N CAUTION however ss cannot delete any columns as this will crash the software 3 Make changes to the file as desired by adding rows deleting rows or adjusting the parameters for each row Some of the items you can adjust are Region e Disorder e Chr e Start e End e Length e Length minus overlap 41 42 CHAPTER 3 User Interface Database of Genomic Variants For example if you would like to more precisely identify the start and stop positions for a particular region you can adjust them in this file You can also add new rows to this file This may be useful if you would like to cross match regions in future samples to regions your lab has tracked in the

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