User`s Manual: SNPStudio_v1.0 [pdf, English]
Contents
1. Weseisa Figure 4 1 Gene Information Table ISTECH SNPStudio 50 v ISTECH BERTAESE Hi na GELIALIE ee Beyond Bioinformatics B SNP List in Gene ACACA 57208415 52305096 52521368 52680395 159900044 57502022 1511650464 lt Figure 4 2 gt SNP Information Table 4 1 2 SNP Location and Gene Structure Creation lt Figure 4 3 gt is the PowerPoint slide showing SNPs and gene structure Gene symbol and gene ID are hyperlinked to the Entrez Gene DB of NCBI The rectangles in the middle of the slide represent SNPs and linked to the gene according to their chromosome positions with arrow line The RNA structures of gene are depicted with exon intron and UTR Green color means UTR and gray color means coding region Please refer to the Chapter 3 3 for detailed control of the visualization C Gene Structure of STAT3 6774 Chr17 Minus Direction 3635 SNP A 2275817 37 719 436 3636 SNP A 2276876 37 723 124 3637 SNP A 1870241 37 725 506 3638 SNP A 2269376 37 733 907 3639 SNP A 1873035 37 745 206 3640 SNP A 2045758 37 748 428 3641 SNP A 4273712 37 751 799 3642 SNP A 2230022 37 767 727 3643 SNP A 2056150 37 768 646 3644 SNP A 4242727 37 783 361 3645 SNP A 4244957 37 787 601 3646 SNP A 1947784 37 795 736 3647 SNP A 1823767 37 796 027 3 Pa NM 139276 2 NP 644805 1 NM 213662 1 NP 998827 1 EA Gait 21 H LI PT i 37 718 865 37 794 039 37 713 869 lt Figure2. ISTECH BGH 88811846 wei Beyond Bioinformatics Figure Legends Figure 1 1 Operating Process of SNPStudio aans i 8 Figure 1 2 Operation Process of Run Ward 9 Figure 2 1 SNPStudio Login Page nennen name nnnnnnnnnnnnnne nennen 13 Figure 2 2 SNPStudio Start Button i 13 Figure 2 3 Application Interface of SNPStudio ennen enn nenneneennen ennn ensen vennen 14 Figure 2 4 Wizard Window of SNPStudio nn annn oen eenneneneennsenseensnenrensenensens sense 14 Figure 2 5 Layout Configuration of OLE Objects errar 15 Fre 26 SNP Data NADU i oro AY E 15 Figure 2 7 Search Multiple Genes by SNPs Location eee 16 Figure 2 8 Search a Specific Gene by Gene Symbol nenne 17 Figure 2 9 Searching and Mapping Status e 17 Figure 2 10 Searching Result naan enreenan eer RL LLE EIRA nn nnnnnnennnnnnnnnnnnnnnn nennen 18 Figure 2 11 List of Gene Relevant Contents i 18 Figure 2 12 Gene Filtering and Gene Selection oneness enen ennnenrennnenrs enen eenen 19 Figure 2 13 Preview of Gene Structures and SNPs Locations nennen ennn 20 Figure 2 14 PowerPoint Template Creaton i 20 Figure 2 15 Completion Message of Run Ward 21 Figure 2 16 List of OLE objects and Summary of Display nennen enn ennn enn enens 21 Figure 2 17 PowerPoint Display e 22 lt Fig
3. ISTECH SNPStudio 19 b ileet smi x ISTE S CH S88118e26 WW Beyond Bioinformatics SNP Studio 2 Gene Structure Preview Normalize Intron 1 MEE Position dbSMP EDR Rate 1 Normalize Exon Adjusted Distance vi 3 NM 00109 NM 02196 n SNP ID Actual Distance Adjusted Distance Chromos Gene Posi NM_0010 NM_0219 Draw S SNP_A 1 Chri T 855 252 rs11247571 ABR 1 74630 Exon244818 Exon25 818 SNP_A 2 Chri rs3263 883 601 rs9910794 ABR 1 74337 ABR 146281 Exon24 525 Intron16 16 Exon25 525 Intron 16 317 847 rs 11855229 ABR 112035 Intron9 2114 Intron10 2114 933 971 rs 17622751 ABR 95911 Intrond 7686 Intron5 7686 953 196 ABR 76686 Intron2 5730 vi nea rs2258130 rs 7207980 rs 12452256 1 006 780 rs2083811 ABR 23102 41 018 374 rs11655015 ABR 11508 Intron3 22074 Intron3 4126 Intron2 28892 Intron2 22933 Intron2 11339 a 971 144 1 000 821 ABA 58738 ABR 29061 Up 12071 Up 41748 Up 47707 Up 59301 co S S ST IST IST EST ST ST S Di 10 SNP A 2 DI Figure 2 13 Preview of Gene Structures and SNPs Locations In this step as shown on the Figure 2 14 you can configure page size and orientation of PowerPoint slide Default configuration is
4. oNPStudio TM Version 1 0 User Manual e y ISTECH Beyond Bioinformatics ISTECH ESTILE G SA na 88811846 wei Beyond Bioinformatics First of all Thank you for purchasing our product This User Manual contains information about SNPStudio version 1 0 It describes notable information in using installation instructions and operating methods Please read the manual before using to manage the program properly This User Manual and the product are protected by the Copyright Law e You can not copy reprint or translate a part or whole User Manual or the product without any written permission of Istech Corp in advance his User Manual and the product may have technical error and this can be adjusted without pre announcement e We will not take any responsibility of the damage caused from using this product beside the purpose and the method described in this User Manual To use this product properly and safely please read this User Manual thoroughly before using Thank you Customer Support Istech Corp is always listening to our customers If you have any questions please feel free to contact us as shown below Thank you Tel No 82 31 903 1155 Fax No 82 31 903 1152 e mail snpstudio istech21 com Website http snp istech21 com snpstudio Technical Support yblee istech21 com ISTECH SNPStudio 2 x ISTECH Beyond Bioinformatics amp e8118 c TA GEELIGA G MA Tabl
5. database of NCBI Line Type Select from off curved line and folded line e Rotate SNP Label about 17 degree in right Select Off not to tilt SNP label B LD Map Adjustment e Display LD Map Select On to show linkage disequilibrium pattern in a reverse triangle e LD Map Color Map Style Six visualization options to change and modify linkage ISTECH SNPStudio 29 v ISTECH BERTAESE Hi AL RESTLESS gei Beyond Bioinformatics disequilibrium pattern Display LD Block Select On to visualize LD Blocks Display Pairwise Tagging SNPs Select On to visualize Tagging SNPs E Haplotype Block Adjustment e Display Haplotypes in LD Block Select On to show the haplotype information estimated in each LD blocks e Display Haplotype Tagging SNPs Select On to show haplotype tagging SNPs estimated in each LD blocks Bl LD Matrix Adjustment Display LD Matrix Select On to show the calculated D and r e Highlight Strong LD D prime Mark SNP pair showing D is bigger than the set up value with yellow color in LD matrix e Highlight Strong LD r2 Mark SNP pair showing r is bigger than the set up value with yellow color in LD Matrix e D Matrix as Excel File Select On to create calculated LD matrix as Excel file ISTECH SNPStudio 30 7 ISTECH GEAIAE RE Mi x 888118e56 wei Beyond Bioinformatics E Prefernece Annotation File Analysis Parameter Visualization Adjustment Template
6. 14 07 060125 call 15 07 060128 call 16 07 060129 call 17 07 060132 call rap N 4 060134 call POC 3 EU x Ll AO MOO OA 0 O O O 10 4 P 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 O EE ig Figure 5 2 Standard Linkage Format Haploview Genotype ISTECH SNPStudio 39 x ISTECH Beyond Bioinformatics E1 Microsoft Excel IGL info m THE BENE HOED EW FUMI Ess eeu A LOD Adobe PDF GERILA G MANN 88811856 WM Lo SACH De r 2 e EE VE CEN SNP A 1970724 SNP A 4235384 SNP A 2192475 SNP A 2118239 SNP A 1917800 SNP_A 4192675 SNP A 2079897 8 SNP_4 2211676 9 SNP_4 4203816 SNP A 2088654 SNP A 4205270 SNP A 1958703 SNP A 2240392 SNP A 4233617 SNP A 4235859 SNP A 22 14994 SNP A 42 16322 20711768 20747696 20825481 20825614 20826146 20871057 20878492 20878522 21049884 21059134 21053244 21181447 21191527 21192042 21251696 21333173 2139458 lm HNIGLG Jis al IEN JES N w DOE KEZ H NUM lt Figure 5 3 gt SNP Annotation Haploview 5 3 International HapMap Genotype Format lt Figure 5 4 gt is an example of International HapMap genotype data For details on file format see http www hapmap org genotypes rs alleles chrom pos strand assembly center protLSID assayLSID panelLSID QCcode NA06985 NA06991 rs9892334 A G chr17 32381183 ncbi B36 affymetrix urn L5ID affyme
7. 7 Genotype of ABI TaqMan 3 Microsoft Excel snp txt BIK m IME BEE ww HUD MAO ERD OOED BW TSH Adobe PDF E X vo E So EM ZAC AI 1 Marker_ID Allele Chr No Chr Pos dbSNP rs 2 ARTS 1E15 88 C G C G 1 1748914 rs10907187 8 ARTS 10 E15 88 C G C G X 74495939 1s5937364 4 ARTS 2 E15 88 G T G T 10 59194086 rs10826109 5 ARTS 3 E15 88 C G C G 3 105657976 rs2673460 6 ARTS 4 E15 88 A C A C 3 7907444 rs495502 7 JARTS 5 E15 88 C T C T 9 33130976 rs7865745 8 ARTS 6 E15 88 C G C G 12 104686299 rs4609683 9 ARTS 7 E15 88 C G C G 6 405901 rs1113387 10 ARTS 8 E15 88 A G A G 8 76366677 rs7840659 11 ARTS 9 E15 88 A T A T 2 129574 rs3007 10 v M 4 M SND Le li xl PIF a ESQ N x OO PU HI NUM Figure 5 8 Marker Information of ABI TaqMan ISTECH SNPStudio 42 ISTECH amp 681185c W i a 888118e58 wei Beyond Bioinformatics 5 6 Result Files from Haploview The output of Haploview program can be used in the SNPStudio lt Figure 5 9 gt is an example of Marker Information Result of Haploview program lt Figure 5 10 gt lt Figure 5 11 gt lt Figure 5 12 gt is each an example of Haplotype Result LD Result and Tagging SNPs of Haploview program 3 Microsoft Excel IGL checkmarker i T E BEE 4700 SEU MAO ED BOED EW SESH Adobe PDF ANN AS Eel SI iE SUN fr mr SS SKJE ER EE RS Lp E 1 f Name Position Obs
8. A4 size and Landscape When the set up is done click Next button then you can see the completion message of run wizard as shown in the Figure 2 15 Click Finish button and it starts the creation of OLE objects based on the previously setup configurations SNPAnalyzer Pro PPT Extension Template Creation Arrange displayed results and create a template PPT Slide Size A4 Paper 210x297 O A3 Paper 297x420 PPT Slide Orientation O Portrait All Objects in One Page Landscape One Object in One Page C More Customization Figure 2 14 PowerPoint Template Creation ISTECH SNPStudio 20 7 ISTECH amp 8811856 SA a 888118 6 AM Beyond Bioinformatics SNPAnalyzer Pro PPT Extension SNPStudio Wizard Completed Setup is sucessfully completed LB If you want to modify the setup option just click Back button Click Finish button to close the SNPStudio Wizard and a PowerPoint file is automatically created Figure 2 15 Completion Message of Run Wizard 2 2 Operation Result Confirmation When the run wizard operation is finished you will have the result as seen on the Figure 2 16 gt On the upper side of the window is the list of created OLE objects of PowerPoint Select one from the list and click Open PPT File button to activate PowerPoint Program Then you will see the visualization result as seen on the Figure 2 17 Click Open Excel File but
9. AD Results 1 Filename ARTS 1 E15 488 C G 3plate PlatelD 4 Assay Type Allelic Discrimination Run DateTime 6 Operator Mal 7 8 Sample Information 9 Marker Setting Information Marker Name Quality Value Threshold 11 ARTS 1 E15 Well Sample Ne Marker Name Allele x Rn Allele Y Rn Call Quality Value Call Type Task Passive Ref 1JA1 ARTS 1 E15 88 C G 3 671 7 998 Both 98 96 Automatic Unknown 3999 844 2 A2 ARTS 1 E15 88 C G 3 088 6 778 Both 99 77 Automatic Unknown 2762 5493 3143 ARTS 1 E15 88 C G 3 184 7 085 Both 99 96 Automatic Unknown 2830 724 4 AA ARTS 1 E15 88 C G 2 939 6 319 Both 98 84 Automatic Unknown 3559 8054 b Ab ARTS 1 E15 88 C G 3 382 2 004 ARTS 1 E15 88 C 99 8 Automatic Unknown 3098 7078 6 A6 ARTS 1 E15 88 C G 3 321 7 351 Both 99 97 Automatic Unknown 2539 6426 FIAT ARTS 1 E15 88 C G 3 461 1 732 ARTS 1 E15 88 C 99 91 Automatic Unknown 3067 0164 8 A8 ARTS 1 E15 88 C G 0 084 8 269 ARTS 1 E15 88 G 99 99 Automatic Unknown 2158 3015 9 Ag ARTS 1 E15 88 C G 0 39 0 524 NTC 100 Automatic NTC 2646 9026 10 A10 ARTS 1 E15 88 C G 0 368 0 49 NTC 100 Automatic NTC 2834 5322 11 411 ARTS 1 E15 88 C G 3 36 7 489 Both 99 91 Automatic Unknown 2282 9639 d 19 A1 ARTS 1 FIR 82 C G 3289 7 3858 Rath 99 AR Automatic 1 Inknawn ORIN 7725 Mi M HA ARTS 1 E15 88 C 47G Splate li gt i Jel2KB TED ww LO P ld ez A RIEN EN CO PO O 100 OI O1 Figure 5
10. G G G G G G G G C G SNP A 1850219 53206 rs1609550 T T ET T T T T T T C T SNP A 2073151 78232 rs8078929 De Jar C C C C A C A C SNP A 2080415 83173 rs4890197 G G G G G G G G G T G T SNP A 1879701 111099 rs8073513 GG G G A G G G A A 14 SNP A 1856953 111223 rs10454094 A T T T T T T T T 15 SNP_A 1791464 113794 rs12947571 G G G G G T G G G T SNP A 1928242 114669 rs4617924 AC C C A C A C AYA SNP_A 2 165788 129225 rs9789059 T T C T C C ET ET 18 SNP A 1788336 129457 rs9788983 A A A A A A A A A we nN chri7 af Poem is ES0 N DO HE RE d In Figure 5 1 SNPAnalyzer Pro Format 5 2 Haploview PLINK Format Figure 5 2 is an example of input data for Haploview and PLINK Program In case of Haploview it is the Standard Linkage Format To locate SNPs on genes annotation file with SNP location information is needed separately along with genotype file E Microsoft Excel IGL ped Do DE BEE Bam NED AAC EAT DOE EW 2278 2402 n x L KSM XI amp ES EA A ER NN NNS ENEN AER EC RE GET EA EZ 07 060101_call o o 0 ilaa AG CC AG TT AG CC CT CG CC TT GG GG AA GG GG GG 07 060104 call 07 060106 call CT O 4 0 0 07 060107 call 0 07 060108 call 0 07 0601 1 1_call 0 07 060113 1 cal O 07 0601 15 call 0 07 0601 16 call 0 07 0601 17 call 0 07 0601 18 call 0 0 0 0 0 0 0 0 O OO O O1 MN A O O O O O O O MH O O O O 07 060 120 call 0 7 060121 call
11. input formats as followings Haploview PLINK Format genotype file and marker information file International HapMap Genotype Format Illumina GoldenGate Format genotype file in matrix format and marker information file ABI TaqMan Format multiple genotype files and one marker information file Result files from Haploview SNPAnalyzer Pro Format one file containing SNP annotation and genotype x Maximum number of SNPs for analysis Over 50 000 x Maximum number of samples for analysis Over 2000 2 2 Run Wizard SNPStudio operates all analyses through run wizard from input data to gene searching SNP mapping and visualization and analysis parameter setup SNP Data Input Gene Boundary Specification Gene Search PowerPoint Creation ListofPPTFlles Template Specification Gene Structure Adjusment dl e e zl Cue Heb Ber Wer ces SE aim r ee m E ee ETE Rm YET NT ir NS Figure 1 2 Operation Process of Run Wizard ISTECH SNPStudio 9 ISTECH BGH ne 88811846 wei Beyond Bioinformatics 1 2 3 Gene Searching and SNP Mapping SNPStudio searches all the genes on which SNPs in the input data are located using chromosome position of SNPs and Ref Seq information provided by NCBI Followings are the information used for gene searching and SNP mapping Official Gene Symbol NCBI Gene ID Start Stop Position of a Gene RNA Count of a Gene RNA ID s Protein ID s Chromosome position
12. searching and SNPs mapping annotation file about genes are required To download new annotation file click Annotation Download tab in the Preference category on the left side of the main interface as you can see in the Figure 3 1 and then a window similar to Figure 3 2 will pops up Click Annotation File tab and select one from the list of annotation files or click Install New Release button Installation of annotation file will proceed after clicking OK button 2 ISTECH AppletViewer Used 44 11M 0 89M DO y Wizard Open PPT amp Preference Support O Exit PPT Create A Open PPT File Open Excel File Adjust Gene Structure Adjust Layout amp Parameters 4 SNPStudio Wizard PowerPoint Excel Pairwise LD Open PPT File SHCLI xIs ABCC3 xls MIRN634 xls Project Management UI Save Project As t Open Project t5 Open Folder VPS53 ls PS53 AATF ppt AATF xls Delete amp Close Project BRCAI ppt BRCAI xls B4GALNT 2 ppt B4GALNT2 xls BRIF 1 ppt BRIP1 xls Preference A BIRCS ppt BIRCS xls BIRC5 8 Analysis Parameter S3 Layout Customization 83 Annotation Download A Gene Structure of VPS53 55275 Chr17 Minus Direction Technical Support fs Product Homepage User Manual Contact enor BE OE 17 818 Ams 219 254 24 1054 54 2649 54 61 SE Mt A4707 mnr 1 MP nr SUP_4 4293630 SMP A 21 ywn A 3832 SMP MAI 3 ss
13. 0 ABR 587308 Up 12071 Intron3 4126 1 000 821 rs12452256 ABR 29061 Up 41748 Intron2 28892 1 006 760 rs2083811 ABR 23102 Up 47707 Intron2 22333 1 018 374 rs 11655015 Up 53301 1 Intron2 11333 KJ 1 KS ES EST EST ST ST Figure 2 19 Gene Structure Adjustment In the Figure 2 16 click Adjust Layout amp Parameters button to change the values of parameters for data analysis and visualization configuration as shown in the Figure 2 207 Detailed usage of the interface will be described in Chapter 3 Parameter Setup 2 Preternece Advanced options Template Adjustment PPT Slide Orientation Landscape Page Size Table Creation e I Te Display Gene Infarmatian C Display SMP Information Gene Structure Creation Display Gene Structure LD Map Adjustment Display LD Map LD Map Color Map Style Display LD Black Display Pairwise Tagging SMPs Figure 2 20 Interface of Parameter Configuration ISTECH SNPStudio 23 5 amp 68118 c SA LAT 6811856 Chapter 3 Parameter Setup ISTECH SNPStudio 24 7 ISTECH amp 881I1856 SA a daas EGET WW Beyond Bioinformatics 3 Parameter Set Up To utilize SNPStudio efficiently it is possible to modify values of parameters for visualization and data analysis 3 1 Gene Annotation Information Download and Installation For gene
14. 3_HYU_CNY370_SNP_tb txt 2127 AS La DME iD CG el ER Ce Ben Name 1 rs10000010 2 rs10000023 3 rs10000030 4 rs1000007 5 rs10000092 6 rs10000121 7 rs1000014 8 rs10000141 9 rs1000016 10 rs10000169 11 rs1000022 12 rs10000272 13 rs10000282 14 rs10000300 15 rs1000031 16 rs1000032 17 rs10000388 18 rs1000040 19 rs1000041 20 rs10000432 21 rs10000435 22 rs10000438 051 re 1NNNNA Index 2 PO Po IND n5 me He i 221218 ISTECH SNPStudio BEE 470 ee Chr 1 1 1 4 4 4 2 4 4 6 4 2 4 3 4 4 4 8 2 4 5 6 4 4 4 GA A n 071213 HYU CNVSTO_SNP tb Eau N X DI O PS lu MAO vo BE USD SHH AC Position 21227772 95952929 103593179 237416793 21504615 157793485 24325037 33810744 235355721 77575270 99259220 189927377 187995996 40161304 44615439 176643771 184885948 23302034 137848410 47206538 70551260 61456357 17588BK1A ED BI OIETDO 1 1 1 1 1 1 1 1 1 1 al sl 1 1 1 1 1 1 1 1 1 1 ECW OT SE El KCNIP 4 BMPRIB NFKB 1 COPS8 GPR125 PDGFC CACNGS CENTD 1 SH3BP4 SHRM CLYBL FLJ36 180 FAT FLJ20273 KAAO427 HOXD13 FLJ12716 UBESA OLIGS ATP 10D UGT241 LPHNS obs Es SE KE i Bis II Bis Bis GG gm IN NETO MII MC META NCI NOT SI II IL II 303 AG IC AA 303 AA 303 D TI tm LL Adobe PDF Gene NM 147183 2 NM 001203 1 NM 0039
15. 4 3 gt SNP Location and Gene Structure ISTECH SNPStudio 34 ISTECH BERTAESE Hi na RESTLESS gei Beyond Bioinformatics 4 1 3 LD Map Adjustment If genotype information is available visualization of the results of linkage disequilibrium LD analysis is implemented automatically as shown in the lt Figure 4 4 gt Strong red color means that there is strong linkage disequilibrium between adjacent SNPs The light blue rectangle on the top of the linkage disequilibrium map is showing tagging SNP LD block is surrounded by thick black line Haplotypes and their frequencies estimated in each LD blocks are tabulated in the bottom of the slide Yellow color is showing major allele of the relevant SNP The haplotype tagging SNP is marked with gray box in the haplotype table For more information please refer to Chapter 3 3 D LD Map E LD Block and Haplotype lt Figure 4 4 gt LD Map and Haplotypes ISTECH SNPStudio 35 GRATES e TA ISTECH Deele 88811846 SM Beyond Bioinformatics 4 1 4 LD Matrix Adjustment Figure 4 5 shows linkage disequilibrium matrix calculated using individuals genotypes As linkage disequilibrium indices D and r values are calculated D values and r values are displayed in the upper triangle and lower triangle separately Complete linkage disequilibrium i e D 1 and r2 1 relationships are colored in yellow as shown in the Figure 4 5 For more detailed informa
16. 62 0 206 0 038 0 016 10 324 0 148 10 114 0 028 0 006 Multiallelic Dprime 0 198 12 BLOCK 3 MARKERS 12 13 14 13 331 0 741 10 523 0 212 113 0 224 10 186 0 038 15 111 0 035 10 029 0 006 Multiallelic Dprime 0 347 17 BLOCK 4 MARKERS 16 17 18 33 0 738 14 0 256 an We nNIGLO H PIAA Se NN DON HRE AIN Ar EEE EH NUM lt Figure 5 10 gt Haplotype Result of Haploview ISTECH SNPStudio 43 x ISTECH Beyond Bioinformatics E Microsoft Excel IGL Id i me BEE EV SNP A 19 t SNP A 22 SNP A 42 4 x 4 x 4 x 4 x 4 x 4 x 4 4 M SNP_A 2 16 SNP_A 2 1 SNP_A 19 SNP_A 41 SNP_A 20 SNP A 22 SNP A 42 SNP_A 19 ERD 3 vo BIZ Le 2E SEN A BIOL ENDO C a SW TSH ZE elt ON M4 gt AIGLE FORRO Le 1 Microsoft Excel IGL tag iB IRE BEE 52 SED EU x OO 23 3 1 captured 15 of 15 alleles at 2 gt 0 8 captured 100 percent of alleles with mean r 2 of 0 986 fusing 12 Tag SNPs in 12 tests Allele 5 SNP_A 1970724 SNP A 4235384 SNP_A 2 192475 8 SNP_A 2118239 9 SNP_A 1917800 10 SNP_A 4192675 11 SNP_A 2079897 SNP A 2211676 SNP A 4203816 SNP A 1958703 15 SNP_A 2240392 16 SNP_A 4233617 SNP A 4235859 18 SNP_A 2214994 SNP_A 4216322 Best Test SNP_A 1970724 SNP A 4235384 SNP_A 2192475 SNP_A 2118239 SNP_A 1917800 S
17. 98 2 NM 006710 4 NM 145290 2 HM 016205 1 NM 006539 2 NM 015230 2 NM 014521 1 NM 020859 1 NM 206808 1 NM 178556 3 NM 005245 3 NM 019027 1 NM 014772 1 NM 000523 2 NM 021942 4 NM 130839 1 NM 175747 2 NM 020453 2 NM 006798 1 NM 015236 3 NM nnRR 9 1 lt Location flanking_5UTR intron flanking_5UTR flanking_5UTR flanking_3UTR flanking_3UTR flanking_3UTR flanking_SUTR flanking_5UTR flanking_5UTR intron flanking_SUTR flanking_5UTR intron intron flanking_5UTR flanking_SUTR flanking_5UTR flanking_3UTR intron intron flanking_5UTR intran SIE SIS LA ra iG AA II AG led SENG gt ae Min MW VE ME 1 dee i dp uei GenomeBuild GeneSymbol LocationRelativeToGene 313145 54687 48339 242030 493482 108729 43799 1933273 169704 7 35955 621734 114015 2047 17694 22007 14209 66813 6619 2740 223b 589077 9 Figure 5 6 BeadStudio SNP Table Golden Gate 4 Coding x 7 ISTECH amp 881I65c W i A EGET wei Beyond Bioinformatics 5 5 ABI TaqMan Format Figure 5 7 is an example of genotype data and Figure 5 8 is an example of SNP information data created from TagMan of ABI EI Microsoft Excel ARTS 1 E15 88 C 47G 3plate txt EIER DE BE 2700 aeb MAO EFT BOB EW SH Adobe PDF 228 24 ANG EX AB WA BENS E SL y B CIA t PSE LL DEEL SIAC AE BLIND F3 EE ES gt gt SE ee eee SDS 2 2
18. Adjustment Table Creation A v Display Gene Information om N wr Gene Structure Creation El cene Subtitles Click Button On the left ta Customize chapter Gene Structure NCBI Hyperlink on the Gene Symbol On wl NCBI Hyperlink on the Gene ID om Height of the Rectangle Representing Exon ix sl Display SNPs over Gene Structure lon lowe Subtitles Click Button On the left to Customize ridx snpid chrpos Surrounding box for SNP Label lon NCBI Hyperlink on the dbSNP rs om Rotate SMP Label about 17 degree in right LEN Figure 3 4 Visualization Parameter Setup ISTECH SNPStudio ol ISTECH GERIKEAG ii nn 6811e26 gei Beyond Bioinformatics Chapter 4 Result Interpretation v ISTECH amp 681185c W i AL RESTLESS gei Beyond Bioinformatics 4 Result of Analysis When the analysis is finished various results will be created as PowerPoint or Excel file 4 1 PowerPoint Result 4 1 1 Table Creation In chapter 3 3 if the Display Gene Information is setup as On as shown in the Figure 3 4 and carry out the analysis basic information of the corresponding gene will be tabulated as shown in the Figure 4 1 If the Display SNP Information is setup as On the information of the SNPs will be tabulated as shown in the Figure 4 2 A Gene Description of ACACA ACACA 31 Mar 27 2008 362 EE NP OMNIS wirmos mem wrmezi wosa EE E wiismosi
19. HET PredHET HWpval Geno FamTrio MendEr MAF Alleles Rating a 1 SNP A 1970724 20711768 0 12 D Tess 1 97 1 0 066 A G 2 SNP_A 4235384 20747696 0 384 0 359 0 5365 95 3 0 235 GA 3 SNP A 2192475 20825481 0 494 0425 0 0509 98 8 0 SUBIT 4 SNP A 2118239 20825614 0 517 0 474 0 3193 100 0 387 A G SNP A 1917800 20826146 0 538 0 499 0 401 98 3 0 476 T C SNP 4 4192675 20871057 0 509 0 482 0 6068 99 4 SNP A 2079897 20878492 0 398 0 388 0 9349 99 4 SNP A 2211676 20878522 0 273 0 253 0 4755 100 SNP A 4203816 21049884 0 342 0 292 0 052 88 4 SNP_A 2088654 21059134 O O 100 SNP_A 4205270 21059244 O 0 100 SNP A 1958703 21181447 0 39 0 384 100 SNP A 2240392 21191527 0 39 0 384 100 14 SNP A 4233617 21192042 0 355 0 347 100 15 SNP A 4235859 21281696 0 128 0 12 100 16 SNP_A 2214994 21393178 0 361 0 376 98 3 17 SNP A 4216322 21394581 0 392 0 392 IGL Jel2KB bl F amp W DO PS RT a Dr 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 A 0 0 0 0 0 0 0 0 0 0 0 06 0 0 0 0 0 4 T z Ej Microsoft Excel IGL haplotype DO i ME BEE 52100 HUD MAO ERD GOIE BW ESH Adobe PDF LX L G led L ss N C xy at Sil AJ v9 HS HS JE SH SLAC AE ZH CND p EE EE 2 1234 0 387 0 136 0 000 3 1412 0 308 10 297 0 005 0 009 E 212 0 173 0 004 0 082 0 085 5 214 0 133 0 039 0 04 0 054 Multiallelic Dprime 0 558 BLOCK 2 MARKERS 6 7 8 8 122 0 590 10 421 0 158 0 014 9 1342 0 2
20. NP 009225 1 2 NM 007295 2 NP_0092261 5 v 3 NM 007296 2 NP 009227 1 25 M 4 NM 007297 2 NP_0092281 17 5 NM 0072982 NP_0092291 2 w E lt Figure 2 8 gt Search a Specific Gene by Gene Symbol lt Figure 2 9 gt shows that the searching and mapping process is running by the Search multiple genes by SNPs location method SNPAnalyzer Pro PPT Extension Sorting SNP List 5412 7656 agains lt Figure 2 9 gt Searching and Mapping Status lt Figure 2 10 gt is the result of searching and mapping The contents in the table in the left side of the window are gene symbol gene ID number of RNAs and number of SNPs located in the E relevant genes You can sort the contents in the table by clicking header of columns Click gt ISTECH SNPStudio 17 v ISTECH BERTAESE Hi m 8881184 amp ee Beyond Bioinformatics button on the right side of the table then you can add or remove the contents in Figure 11 gt to the table by checking or unchecking SNPAnalyzer Pro PPT Extension Gene Selection Official Gene Symbol LOC653051 NCBI Gene ID 653051 Chromosome 17 Total SNP Count 7656 M d SNP 3 7656 5 Upstream Region 5 000 pou d 3 Downstream Region 5 000 Total mapped SNP 4193 7656 Total Genes 962 1468 RNA ip No RNAID Protein ID BIER 1 AM 925795 2 XP 930888 D Fie No Gene NCBI ae RNA s Symbol GenelD Count LOC653 653 061 Sel
21. NP rs This is the fixed SNP ID in the NCBI dbSNP database It is a mandatory reserved word e The Remaining Columns These are individual IDs If there is no genotype information there is no need to describe B The Second Row The First Column Sample_Type This is for the next version of SNPStudio It is a mandatory reserved word e The Second The Fourth Column It is marked with It is a mandatory reserved word e The Remaining Columns These are for the next version of SNPStudio and marked in 0 or 1 If there is no genotype information these are not necessary B The Remaining Rows These are the actual values corresponding to each columns of the first row ISTECH SNPStudio 38 z ISTECH 8884166 Hit A 888118e48 wei Beyond Bioinformatics Ej Microsoft Excel chri 7 geno SUE Bae 470 Aed MAO ED HOED BW SCH Adobe PDF LL j e Di x at St l Y 62 HS SHH SIAC AE BLIND J a 1 Marker_ID Chr No Chr Pos dbSNP_rs VFOOO66 Call VF00126 Cal VFO0130 Cal VFO0141 Call VFOO161 Call VFO0164 Call Sample Type 0 0 O 0 0 0 SNP_A 2098700 6888 rs1106175 A G AVA AVA AA AYA AVA 4 SNP_A 1837999 18901 rs8064924 C T T T T T T T DAT T T SNP_A 222 1204 34276 rs3794811 C G G G G G G G G G G G 6 SNP_A 4304383 38761 157224313 C C C C C G C C C C C C SNP A 1941777 43474 rs4890199 G G G G G G G G G G G G 8 SNP_A 4218752 46286 rs4890173 G G G G G G G G G G G G 9 SNP A 1780843 52467 rs7503116 G G
22. NP_A 4192675 SNP_A 2079897 SNP_A 22 11676 SNP_A 42038 16 SNP_A 1958703 SNP_A 1958703 SNP_A 1958703 SNP_A 4235859 SNP_A 2214994 SNP_A 2214994 vog HS EM MIC 2 west M4 gt Ah IGLE Jgl2K B ISTECH SNPStudio ESD Ne LI CO ES l Adobe PDF 8891184 G si 88811856 gei EIER E led A4 io Ir 113713 113846 114378 159289 166724 166754 338116 469679 479759 480274 OD 77918 78450 123361 130796 130826 302188 433751 Figure 5 12 Tagging SNPs of Haploview 44
23. d td Open PPT Fila Ci fne Project da Aben SNP Sandie I M SNP mbob a cnw e program nach mansiones SSP gorun guo a Po er DLE objecr web aid penome and pene imenges Sx Pede kat folu emnes e Super een formen Dnreznamond Hapdlag formar Heploves form enes Gender arc forest ADI Tip Man borum and cur p prester SN P Aaa Pro format Alte comua ci created as DLE objet and eid no s r sangere fo eral erun eent veie Powe Eog Veu SNP aelured maler mt HOED i SS Prado GMP seppe LD torg hapkorupe esoe sm mer SNPs serion e Wied of SP Senden maker vom NT related eiken ear sad be Technical app fp Prout Hepie iT User Mesa 7 Contaeci TERWUT H LI TIGRE Run Wizard ENP ho operates all anal through a run wid Folkeeing in am tapir of pong prose Lead Se pesci ak iori ERP Duda ing Gere Mere y Specie Sien Gorm Search o o FCT m x lt Figure 2 3 gt Application Interface of SNPStudio 2 1 Run Wizard SNPStudio always starts from run wizard Click SNPStudio Wizard on the left side of the application interface or click the hot key Wizard on the top of the interface then a window of run wizard like Figure 2 4 pops up Click Start button to go to the next step EA SNPAnalyzer Pro PPT Extension E3 Se SNPStudio wizard Start MISTECH Start SNPStudio Wizard 2133 4 IRA iii SNP Studio is a software program that visualizes SNPs and gene EN a WN ier mir st
24. e of Contents Copyright 2008 ISTECH INC rs 1 1 MAMOG UCHOD iest nenten dee 8 1 1 S UI ANY RE RE 8 1 2 FNS n S 9 1 2 1 Je V dB Kn En 9 1 2 2 SE MA 9 12 9 Gene Searching and SNP Mapping nn ennen eenenseneennsen 10 1 2 4 Linkage Disequilibrium Analysis eneneneneeenenr eenen ennnnenen vens 10 1 2 5 Creation of OLE Object of PowerPoOINt ene nen eneen enen 10 1 3 System Heourement iii 11 2 Login and Start SNPStudio i 13 2 1 RAVN 14 2 2 Operation Result Confirmation i 21 o PM ENN 25 Sa ha Gene Annotation Information Download and Installation 25 3 2 Analysis Algorithm Parameter Setup eenn 26 3 3 Visualization Parameter Setup ii 28 4 Lg ENEE 33 4 1 PowerPoint Result 33 4 1 1 Table Creation EEE RR O UG RR OR 33 4 1 2 SNP Location and Gene Structure Creation eneen 34 4 1 3 LO Map UE EE 35 4 1 4 LD Matrix Adjustment annen AAA HLR nennen nennen nnn nenas 36 gt AMDT Aber IHN 38 S SNP Analyzer Pro en E 38 5 2 Haploview PLINK Format esses nme 39 5 9 International HapMap Genotype Format ann ii 40 5 4 Illumina Golden Gate Format 41 ISTECH SNPStudio 3 x ISTECH amp 681185c W i RT ema 88811846 Mi 2 9 ABI TagMan Format 5 6 Result Files from Haploview ISTECH SNPStudio 4
25. ect genes to be displayed in PowerPoint Gene Information RPH3AL 9 501 elected Structures LOC400 400 566 VPS53 55275 FAMSTA 79 850 GEMIN4 50 528 ClTod25 51 031 RNMTLI 55 178 4 AISHA ca rn Gene Sym Gene ID Dl co 01 w MI lt Figure 2 10 gt Searching Result Gene symbol NCBI i GenelD Start Pos Stop Fos v SNP HN A M Count Gene Orient column harizontal amp croll column nac kAll lt Figure 2 11 gt List of Gene Relevant Contents As seen on the Figure 2 12 gt input AB on the empty text box right below the Gene Information title then genes with gene symbols starting with AB only will be listed in the table Select the gene that you want to create as OLE object then the relevant RNA IDs and protein IDs will be listed on the right side of the window Select RNAs in the list using check box then these will be created as OLE objects of PowerPoint If you select gene in the list titled as Selected Structures and uncheck RNAs in the list titled as RNA ID then you cancel the creation of OLE objects about the relevant gene You want all the genes listed in the left table to ISTECH SNPStudio 18 ISTECH ETC MM dn Ben Mi Beyond Bioinformatics be created as OLE objects just click Select All button Click None to cancel the checking status Preview button enables use
26. enone WEE m A Op MEE 172 AE DA i HEN Figure 2 17 PowerPoint Display Ej Microsoft Excel CARD14 xIs E ERD HOED KW ESH Adobe PDF wE E r I E IS x al Si d b CARS USA SI SLAC SE ELD ME 0x9 0613 0607 0135 05283 0 1843 M 0 144 0093 011996 0925 E 5 s 6 IS era oM Pairwise D el P Figure 2 18 Excel Display In the Figure 2 16 click Adjust Gene Structure button then you will have the interface as seen in the Figure 2 19 Detailed control options and usage are identical as describe ahead ISTECH SNPStudio 22 7 ISTECH 86811626 SNR u 888118e58 Mi Beyond Bioinformatics SNP Studio 7 v Gene Structure Preview Exon Intron Rate dE Normalize Exon Normalize Intron LE Actual Distance 3 NM 00109 DI wn Br N Adjusted Distance NM 02196 10 00000 Dt OD DIA E 34 D H 12 3 4 5 6 7 89 10 Na SMP ID Chromos Position dbSNP Gene Posi NM 0010 NM_0219 Draw S 1 SNP ai Chri 855 252 rs11247571 ABR 174630 Exon24 818 Exon25 818 865 545 153263 ABR 174337 Exon24 525 Exon25 525 883 601 rs9910794 ABR 146281 Intron16 16 Intron17 16 v 917 847 rs11655229 ABR 112035 Intron9 2114 IntroniO 2114 v 933 971 rs17622751 ABR 95911 Intrond 7686 Intronb 760b 953 196 rs2258130 ABR 76686 Intron2 5730 Intron3 22074 971 144 rs 720738
27. id perlegen hapmap org Protocol Genotyping rs7222936 C T chr17 32393438 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs8069751 A G chr17 32394215 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping Figure 5 4 Internal HapMap Genotype Format ISTECH SNPStudio 40 EGATIGA G smi EGET wei x ISTECH Beyond Bioinformatics 5 4 Illumina Golden Gate Format It is able to use GoldenGate format data of Illumina as input file The Figure 5 5 is an example of the genotype result data extracted using matrix format in the BeadStudio program provided from Illumina The Figure 5 6 is an example of the result data by saving SNP Report Table displayed in the interface of BeadStudio program Ei Microsoli Excel 071213 HWLI Genolype Dalai 217 HE Ie LEE sm gen vam ERD AMED ew EH Adobe POF fe M EI BI ET EIS TI HT IJ ET L MUN PO R 1 Header BST Verdon Pracessng Date Content Num SHES B Total SNPs Num samples Total samples Data 10 11 rs10000010 rs 10000028 rs 10000080 14 rs1000007 rs 100092 rs10000121 rs 1000014 18 rs10000141 19 rs1ODDD 18 20 rs 10000168 21 rs 1000022 re 10000272 re 10000282 re 10000300 3 1 14 1128 2007 2 31 FM Human PWS file C bpm 30404 30404 Gata ae 32 Hak MOTEL Hl GenoteneDate dl Lal CC IG ais IC II iG AA IC LL qi 303 CC E Microsoft Excel 07121
28. igure 5 8 Marker Information of ABI TaqMan ennn ennnensennnenen ven vennen 42 Figure 5 9 Marker Information Result of Haploview nennen nennen 43 Figure 5 10 Haplotype Result of Haploview nun i 43 Figure 5 11 gt LD Result of Haploview A 44 Figure 5 12 Tagging SNPs of Haploview un unn sun ensenense ene ennsensennenenreenenenseneennnen 44 ISTECH SNPStudio 6 amp 681X846 MANN DIES GEELIGA G MA Chapter 1 Introduction ISTECH SNPStudio d z ISTECH GERALA G W i EGET EE WM Beyond Bioinformatics 1 Introduction 1 1 Summary SNPStudio is a software program which transforms SNP information into a PowerPoint OLE object with associated genome and gene structures And if there are genotype information oNPStudio automatically performs various analyses like calculation of linkage disequilibrium LD among adjacent SNPs construction of LD blocks and haplotype estimation within the constructed LD block It is possible to analyze up to tens of thousands SNPs and whole analysis procedure is controlled in detail through SNPStudio run wizard The analyzed results are automatically created into OLE objects and easy to edit manipulate for visual enhancements within PowerPoint Figure 1 1 Operating Process of SNPStudio ISTECH SNPStudio 8 ISTECH amp 68118c W i RESTLESS gei Beyond Bioinformatics 1 2 Main Features 1 2 1 Input Data Type SNPStudio supports various
29. marker information data Genotype File SNPStudio supports three input file Formats 10K test 22 geno SNPAnalyzer Pro Format one file with SNP annotation and genotype O Haploview PLINK Format genotype files marker information file Content Marker 148 snp SNP_A 2113228 SNP_A 2125836 O Illumina GoldenGate Format Sample 172 samples genotype file in matrix format marker information file 07 060101_call 04 060309_cal1 O ABI TaqMan Format Chree 15 607 742 46 466 315 148 snp genotype file marker information File International HapMap Format SNPStudio can do visualization work using following data O Result files from Haploview LD block amp haplotype lt Figure 2 6 gt SNP Data Input ISTECH SNPStudio 19 ISTECH BERTAESE Hi na RESTLESS gei Beyond Bioinformatics On this step it is possible to set up searching method of the genes in addition to SNP mapping Followings show details of the searching method B Search multiple genes by SNPs location search for all the genes on which SNPs in the input data are located B Search a specific gene by gene symbol search for a specific gene using gene symbol or NCBI gene ID The default values of 5 Upstream Region and 3 Downstream Region are 5 000 bp and these mean that the upstream and downstream boundaries of the genes on which SNPs in the input data are located are 5 000 base pairs each Figu
30. nd tagging SAPs selection S PRU a Wired of SNPStudio makes your SNP related tasks easy end fun L Va 7 8 r XJ Sp Go Figure 2 1 SNPStudio Login Page After login scroll down the page and click Start Application button as shown in Figure 2 2 then it triggers application interface of SNPStudio as shown in Figure 2 3 J 5HP Studia Stan Page Microsoft Internet Explorer TE BEE wo BURN FAM TARH a e e CH Lk T a d 9e 9 HAG Can Ze aaen e 25 EU WD3 vU infin ang istech cam snpstadia D Tel 82 31 903 1155 FAA 52 31 903 1152 E mail enpetuchotristech com Lee Yong Bock Hompase ber mechi 1 com r nine Purchase Jaw tech com Eos Wi Start Application Program Message License Period English e Never expres Feature Table Feature Table e Ihmina Genonpe e ABI Taglian Internabonal Hap Wap Hapleview Input Data Output Data Compatible e PLINK Input Data Compatible SKPAnalzer ProTM il Thana i PowerPoint File a byr d Fila Figure 2 2 SNPStudio Start Button ISTECH SNPStudio 13 v ISTECH amp 8811856 SA RIS TECH 86811848 Ai Beyond Bioinformatics On the upper side in the application interface there are hot keys like Wizard Open PPT Preference Support and Exit On the left side of the interface there are menus required to operate and manage the program a EMP Emelie Wirar
31. nstalled with SNP Analyzer Pro MicroSoft PowerPoint 2003 or higher ISTECH SNPStudio 1l ISTECH GERIKEAG nn 6811e56 gei Beyond Bioinformatics SNPStudio Operation 7 ISTECH 86814646 W HL re 888118 6 AM Beyond Bioinformatics 2 Login and Start SNPStudio SNPStudio is a software program which operates on web browser after downloading execution file Figure 2 1 shows the homepage of SNPStudio and the application program can be implemented after login with your ID and password J SNP S1udio Stan Page Microsoft Internet Explorer HIE BEE 4700 BARN EHT TASHI Gwe O iz f s yew 2 5 3 gs DL hipstere latech 1 cam snpstadioy 1 0 v EI us PASSWD ERP Studio E About SNPSimdio IM TutorTa Input File Format Gi SNPStadio is a software program which transforms SNP information into a PowerPoint OLE object with associated un urn genome and gene structures SNP Studio has followmg features 3 Other Product A FE Supports various input formats Intemational HnpM ap format Hapleiev format Illumina GoldenGate format ABI SNPADA Yzer O Fr tnd TaqMan format and cur proprietary SNPAnalyzer Pro format Sue arate er_l Frege Allthe output is erented p OLE object and easy to edit manipulate for visual enhancements within PowerPoint a Varios SWP relared analyses are automated m SAP neo SNP mappe LD blocking baplonpe estenation a
32. of SNPs 1 2 4 Linkage Disequilibrium Analysis If the input data contains individuals genotype information SNPStudio automatically performs linkage disequilibrium analyses The contents of the linkage disequilibrium analysis are as follows Measuring Linkage Disequilibrium between Adjacent SNPs E Constructing Linkage Disequilibrium Blocks LD Blocks E Estimating Haplotypes in Each LD Blocks E Pairwise Tagging SNPs Selection E Haplotype Tagging SNPs Selection 1 2 5 Creation of OLE Object of PowerPoint All the analyzed results are created as OLE objects of PowerPoint of MicroSoft and easy to edit manipulate for visual enhancements within PowerPoint Followings are the types of OLE objecis E SNP Information e SNP ID dbSNP Zrs Chromosomal Position and etc E Gene Information Gene Symbol NCBI Gene ID Gene Orientation mRNA ID Protein ID e Exon Intron UTR e Etc ISTECH SNPStudio 10 ISTECH BEATE SE Hi RESTLESS gei Beyond Bioinformatics B Linkage Disequilibrium Information e Linkage Disequilibrium Map Block Matrix e Haplotypes in LD Blocks Pairwise Tagging SNPs e Haplotype Tagging SNPs e Etc 1 3 System Requirement u Minimum Specifications OS Microsoft Windows 2000 XP Vista System internet connection required e CPU Pentium 4 2 4GHz or higher e RAM 1GB or more Web Browser Internet Explorer 6 0 or higher E Required Application Program e J2SE Runtime Environment 5 0 or higher i
33. on Select whether landscape or portrait e Page Size Select whether A3 or A4 size m Table Creation e Display Gene Information Select On to create the gene information related table e Display SNP Information Select On to create the SNP information related table E Gene Structure Creation Display Gene Structure Select On to visualize gene structure e Gene Subtitles It is able to edit add remove labels related to the gene using following parameters chapter identifier of OLE objects default values are A B C genesymk gene symbol geneid NCBI gene ID chrno chromosome number geneorient gene orientation NCBI Hyperlink on the Gene Symbol Select On to create hyperlink to Entrez Gene DB of NCBI NCBI Hyperlink on the Gene ID Select On to create hyperlink to Entrez Gene DB of NCBI e Height of the Rectangle Representing Exon Set up the height of exon compared to the default value ex 4X means four times the default height e Display SNPs over Gene Structure Select On to visualize SNPs locations with the relevant gene e SNP Subtitles It is able to edit add remove labels related to the SNPs using following parameters ridx serial number of SNP starting from 1 snpid dbSNP rs ID chrposk position on the chromosome e Surrounding box for SNP Label Select On to draw box surrounding SNP label e NCBI Hyperlink on the dbSNP rs Select On to create hyperlink to dbSNP
34. r to preview and modify the SNPs locations and gene structures in a separate window as shown in lt Figure 2 13 gt Click Next button to go the next step SNPAnalyzer Pro PPT Extension Gene Selection Official Gene Symbol ABR MOBI Gene ID zu CRWT 7 Total SNP Count 7656 ul d SNF 10 7656 5 Upstream Region 5 000 Fe 3 Downstream Region 5 000 Total mapped SNP 4195 7656 Total Genes 882 1458 RNA ID DNA ID Protein ID Select genes to be displayed in PowerPoint Gene Information Mm 001092 3 MP ME 2 MM_021962 2 NP _068 ie ii ABR e Figure 2 12 Gene Filtering and Gene Selection In the Figure 2 13 you can modify SNPs locations and gene structures using four configuration methods B Exon Intron Rate Control the ratio of the length of exon and intron B Normalize Exon Control the relative lengths of exons regarding other exons lengths B Normalize Intron Control the relative lengths of exons regarding other introns lengths m Actual Distance Adjusted Distance If you do not want to control the ratio of the length of Exon Intron select Actual Distance option in the list of the right side of the window Default value is Adjusted Distance Preview of gene structures and locations of SNPs are displayed right below the control panel You can add or remove SNPs for visualization using check box Click Yes button to apply the modified configuration and proceed to the next step
35. re 2 18 gt Excel RTE 22 Figure 2 19 Gene Structure Adjustment oan oon eenn enn eeneneeneennsenrsennenreerenensensennnen 23 Figure 2 20 gt Interface of Parameter Configuration nnens ene ennnnenrennnenr ennen 23 Figure 3 1 gt Main Interface of SNPStudio aan LLYR enn eenenernennnsenrvennenrserenenseneennnen 25 Figure 3 2 Annotation File Download nnn i 26 Figure 3 3 Analysis Parameter Gen 28 Figure 3 4 Visualization Parameter Setup nn ansnnnnennennnsenneennsenrennnenrn ven ennnsens ennen 31 Figure 4 1 Gene Information Table i 33 Figure 4 2 SNP Information Table eenen mnes 34 Figure 4 3 SNP Location and Gene Giruchure eene 34 Figure 4 4 LD Map and Haplotypes ei 35 lt FIgure 4 5 gt LD Mat Ae sw ara a EA anne na OCH daria ia 36 Figure 5 1 SNPAnalyzer Pro Format enn ennn enen eneneerenenenenvensennneerenensenesennen 39 Figure 5 2 Standard Linkage Format Haploview Genotype annen LL nennen 39 ISTECH SNPStudio 5 x ISTECH BERTAESE Hi ert eng 888118 6 wg Figure 5 3 SNP Annotation Haploview eee LL LLE LU LLC LL FY FFF F Yno 40 Figure 5 4 Internal HapMap Genotype Format errar 40 Figure 5 5 GoldenGate Genotype Format ii 41 Figure 5 6 BeadStudio SNP Table Golden Gater errar 41 Figure 5 7 Genotype of ABI TaqMan naan onsen ennn enenerrenensnenvensennneerenenseneennnen 42 F
36. re 2 7 is the interface to search for all the genes on which SNPs in the input data are located Click Next button to go to next step SNPAnalyzer Pro PPT Extension SNP To Gene Mapping Search genes and RNAs on which input SNPs are located using chromosome positions of SNPs Search multiple genes by SNPs location Annotation Library Detail Release Data SEP 21 2007 Entrez Release 35 2 dbSNP Release N A Content O Search a specific gene by gene symbol F Upstream Region 000 bp 3 Downstream R 5 000 bp Figure 2 7 Search Multiple Genes by SNPs Location Figure 2 8 is the interface to search a specific gene using gene symbol or NCBI gene ID Input gene symbol or NCBI gene ID and click Search button to see the table with the associated information such as RNAs and proteins Select RNAs using check box to be created as OLE objects Click Next button to go to next step ISTECH SNPStudio 16 V ISTECH EGALIG mi a 888118 6 gei Beyond Bioinformatics 2 SNPAnalyzer Pro PPT Extension SNP To Gene Mapping Search genes and RNAs on which input SNPs are located using chromosome positions of SNPs Gene Symbol NCBI GeneID Search O Search multiple genes by SNPs location Search a specific gene by gene symbol NCBI Gene ID 672 S Upstream Reglon bp Total SNP Count 7656 3 Downstream H bp Mapped SNP No RN Protein Exon NM 007294 2
37. rg Equilibrium test if the calculated p value is smaller than the fixed value corresponding SNP will be excluded from the analysis ISTECH SNPStudio 26 ISTECH BGH na GELIALIE wei Beyond Bioinformatics e HWE p value multiple correction Apply Bonferroni multiple test correction m Tagging SNPs Selection e Minimum allele frequency threshold If the observed allele frequency is smaller than the fixed value corresponding SNP is removed from the list of tagging SNPs e r2 threshold If the square of correlation coefficient r between adjacent SNPs is smaller than the fixed value corresponding SNPs are removed from the list of tagging SNPs E LD Blocking Gabriel s method Lower D e Upper D e Strong LD fraction Minor allele frequency e Maximum Segment Limit e Four Gamete Rule Min Haplotype Frequency x Please refer to the related thesis Gabriel et al The structure of Haplotype blocks in the human genome Science 2002 296 5576 2225 2229 for detailed description of the parameters E Haplotype Tagging SNPs Selection e Entropy Reduction x Please refer to the related thesis Avi Itzhak et al Selection of minimum subsets of single nucleotide polymorphisms to capture Haplotype block diversity Pac Symp Biocomput 2003 466 477 for detailed description of the parameters The parameters used for linkage disequilibrium analysis can be set up by clicking Analysis Parameter tab from the Preference ca
38. ructure upon which the SNPs are mapped using physical location Ex of the SNP on the human genome start 4 SNPStudio Install Wizard provides diverse control options for _ About visualization and user can get a proper result by combining them The final results are PowerPoint files of MicroSoft Inc Please click Start button to continue mm Figure 2 4 Wizard Window of SNPStudio ISTECH SNPStudio 14 x ISTECH GERE MN Beyond Bioinformatics sol 6 6 AA aal On this step it is possible to select layout configuration from four different options as shown in the Figure 2 5 Selecting each item you can see the example result on the right side of the interface Click Next button to go to the next step 2 SNPAnalyzer Pro PPT Extension Object Selection Select one of the layouts to be displayed in PowerPoint O SNP amp Gene Structure O SNP amp Gene Structure with LD Map amp Haplotype O SNP amp Gene Structure with LD Matrix SNP amp Gene Structure with LD Map amp Haplotype amp LD Matrix Figure 2 5 Layout Configuration of OLE Objects On this step you can select one input data format among six different formats as shown in the Figure 2 6 Select relevant data format and click Open button to input your data When the data input is done click Next button to go to next step SNPAnalyzer Pro PPT Extension SNP Data Input Input SNP genotype and
39. ss NS iN unpre me LI N NY Figure 3 1 Main Interface of SNPStudio ISTECH SNPStudio 25 v ISTECH amp 68118c W i na RESTLESS gei Beyond Bioinformatics Prefernece Annotation Download NCBI Genome B SEP 21 2007 Release 3 v lt Figure 3 2 gt Annotation File Download 3 2 Analysis Algorithm Parameter Setup It will automatically operate the Data Preprocessing and Linkage Disequilibrium Analysis between SNP if there is an Individual Genotype Information in the input data Followings are the explanations of the Parameter used for the analysis E Preprocessing e Replace missing genotype with Replace individual genotype without observed value with one from three possible genotypes major homozygous genotype heterozygous genotype minor homozygous genotype e Flag SNP with missing genotype frequency over Among observed genotypes if the ratio of missing genotype is bigger than the fixed value corresponding SNP will be excluded from the analysis e Flag sample with missing genotype frequency over Among observed genotypes if the ratio of missing genotype is bigger than the fixed value corresponding sample will be excluded from the analysis e Flag SNP with minor allele frequency below If the observed allele frequency is smaller than the fixed value corresponding SNP will be excluded from the analysis e Flag SNP by HWE test p value below From the result of Hardy Weinbe
40. tegory on the left side of the interface as seen on the Figure 3 1 If you want to control hidden parameters check Advanced options as shown in the Figure 3 3 ISTECH SNPStudio 27 7 ISTECH G6FIAE SE W i AD 8881184 amp ee Beyond Bioinformatics 2 Prefernece Annotation File Analysis Parameter Visualization Adjustment Preprocessing Replace Missing Genotype with Which Genotype Restore Missing De ZI cen Flag SMP with missing genatype frequency aver cr Flag sample with missing genotype frequency over a cea c Flag SMP with minor allele frequency below Flag SNP by HWE test p value below HWE nvalue multple correction Tagging SNP Selection c c LT Min Haplotype Frequency Threshold Lem CO r Threshold LD Blocking ea Lower D 5 LO ea Upper D Strong LD Fraction Minor allele frequency lo m Maximum Segment Limit 5 Four Gamete Rule Min Haplotype Frequency Haplotype Tagging SMP Selection Entropy Reduction lt Figure 3 3 gt Analysis Parameter Setup 3 3 Visualization Parameter Setup It is possible to set up parameters used for visualization of SNP and the gene structure with SNP mapping LD map and etc as shown in the lt Figure 3 4 gt E Template Adjustment ISTECH SNPStudio 28 ISTECH ESTILE G SA RESTLESS Gd Beyond Bioinformatics e PPT Slide Orientati
41. tion please refer to Chapter 3 3 F LD Matrix 2 3 5 5 7 9 10 11 1 0 972 0 991 1 110 973 1 1 0 991 0 963 0 991 0 01 0 011 0 01 0 566 0 555 0 569 0 789 0 763 0 79 0 79 0 763 0 791 0 793 0 794 0 991 0 99 0 01 1 0 578 0 01 0 492 0 492 0 559 0 492 0 485 0 547 0 475 0 475 0 541 0 475 0 467 0 529 0 496 0 496 0 562 0 496 0 488 0 55 0 503 0 044 0 503 0 044 0 57 0 012 0503 0 044 0 496 0 041 0 558 0 012 0 906 0 906 0 906 0 903 0 982 0 991 0 892 0 391 0 99 0 991 0 892 0 889 0 01 0 011 0 01 0 044 0 044 0 041 0 012 001 o lt Figure 4 5 gt LD Matrix ISTECH SNPStudio 36 amp 681X846 MANN DIES GEELIGA G MA Beyond Bioinformatics Chapter 5 Input Data ISTECH SNPStudio 37 ISTECH BGH 88811846 wei Beyond Bioinformatics o Input Data Format 5 1 SNP Analyzer Pro Format This is the tab delimited text file format The first line and the second line are used as headers which are followed by the SNP information and individual genotypes Figure 5 1 shows an example and details are as follows B The First Row The First Column Marker ID This is to classify SNP It is a mandatory reserved word The Second Column Chr No This is the chromosome number of SNP It is a mandatory reserved word The Third Column Chr Pos This is the location of SNP on the chromosome It is a mandatory reserved word The Fourth Column dbS
42. ton to activate Excel program and you will have the linkage disequilibrium analysis result D and R2 as you can see on the Figure 2 18 IS TECH Appleren lined S51 2234 IM Wizard Open PPT X Preierence 7 Support Exit Open PPT Fie Adios Gene Suche II Agus Layout amp Parameters PPT Croato D SMPPuda Wired amp Oran PPT Fila Excel Pairwise LD SHCLI npa SMCLI sig SC AES C3 ppa AES C3 che ABELS WISEN xls MANGE TU Save Project Ad le Open Project 5 Open Folder MX Delete amp Clete Fra HS EP ppi ALE ppi Kai e Technical Support fi Product Homepage LEI User Maan E Contact Gene Structure ol CARD 4 19092 Chr17 E LO Map Fatz ur Bia nm rm Fun Sep Elum PFFF ini FG Pet bs En pisi D LU Matrix i j H E SLI Drs 04642 VEE JJ e 0 108 Sau OACL DIM L L gor 5 194 Fi Fei Ere nmn i giii GORT ou pars FL 0254 0009 0 21 DIEP Oath BOB OB DIM 0 408 i 0 179 6 012 009 Gs 0 143 lt Figure 2 16 gt List of OLE objects and Summary of Display ISTECH SNPStudio zi ISTECH 86841626 G i RISTECH Ls Beyond Bioinformatics ME ode a ES ae A Gene Structure of CARD14 720927 Chr17 Lg DAP 5 TP TEE SNP DD AUR FETTE GE EETL FET ETH FIA SNP PIE 75 771 554 va Kr SAP A RS RATIO AG GH AAPG Pb TEE SHE AE PR TREE RIT lee Steet IM GOIS E VER P
43. trix hapmap org Protocol genotyp rs8077789 C G chr17 32383839 ncbi B36 imsut riken urn lsid imsut riken hapmap org Protocol genot rsi0908286 A C chr17 32383961 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs11650575 A C chr17 32384701 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs8067751 A C chri 32384744 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs12051731 C G chr17 32384818 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs4408589 C G chr17 32385876 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs7218038 A C chr17 32389341 ncbi B36 affymetrix urn LSID affymetrix hapmap org Protocol genotyp rs6607358 A G chr17 32389367 ncbi B36 illumina urn L5ID illumina hapmap org Protocol Infinium ge rs11871752 C T chr17 32389923 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs9895466 A G chr17 32390954 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs9972935 A G chr17 32391264 ncbi B36 imsut riken urn lsid imsut riken hapmap org Protocol genot rs9893214 C T chr17 32392031 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rsii871466 A G chr17 32392482 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs4427852 A T chri 32392660 ncbi B36 perlegen urn lsid perlegen hapmap org Protocol Genotyping rs12942174 A G chri 32393071 ncbi B36 perlegen urn ls
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