Genotyping Technology How to Analyze Your Own Genome Fall 2013

Transcription

1 Genotyping Technology How to nalyze Your Own Genome Fall 2013

2 HapMap Project Phase 1 Phase 2 Phase 3 Samples & POP panels Genotyping centers Unique QC+ SNPs 269 samples (4 populations) HapMap International Consortium Reference Nature (2005) 437:p samples (4 populations) Perlegen 1.1 M 3.8 M (phase I+II) Nature (2007) 449:p851 1,115 samples (11 populations) road & Sanger 1.6 M (ffy 6.0 & Illumina 1M) Draft Rel. 1 (May 2008)

3 Phase 3 Samples label population sample # samples QC+ Draft 1 SW* frican ancestry in Southwest US CEU* Utah residents with Northern and Western European ancestry from the CEPH collection CH Han Chinese in eijing, China CHD Chinese in Metropolitan Denver, Colorado GIH Gujarati Indians in Houston, Texas JPT Japanese in Tokyo, Japan LWK Luhya in Webuye, Kenya MEX* Mexican ancestry in Los ngeles, California MKK* Maasai in Kinyawa, Kenya TSI Toscans in Italy YRI* Yoruba in Ibadan, Nigeria ,301 1,115 * Population is made of family trios

4 HapMap rowser 1a. Go to 1b. Select HapMap phase 3

5 Overview Genotyping technology - SNPs and copy number variaeons Processing the data from genotyping assays - Linkage disequilibrium - Haplotype inference, phasing - Tag SNP seleceon Preliminary analysis of HapMap data

6 SNP Genotyping with SNP rray SNP arrays make use of the biochemical principle that nucleoede bases bind to their complementary partners ( binds to T, C binds to G) n array of oligonucleoede sequences is laid across the surface of the chip. The sample s DN is amplified, and hybridized to the array. The array is scanned to quanefy the relaeve amount of sample bound to each feature. For SNPs, there is a pair of probes: one for each of the alleles. Widely used SNP array technology ffymetrix vs. Illumina SNP arrays

7 ffymetrix GeneChip Probe rray

8 SNP rray Technology: ffymetrix rray The fragment of DN harboring an /C SNP to be interrogated by the probes 25- mer probes for both alleles The locaeon of the SNP locus varies from probe to probe 25- mer (25 nucleoedes) The DN binds to both probes regardless of the allele it carries, but it does so more efficiently when it is complementary to all 25 bases (bright yellow) rather than mismatching the SNP site (dimmer yellow). This impeded binding manifests itself in a dimmer signal.

9 SNP rray Technology: Illumina rray The fragment of DN harboring an /C SNP to be interrogated by the probes \ached to each Illumina bead is a 50- mer sequence complementary to the sequence adjacent to the SNP site. The single- base extension (T or G) that is complementary to the allele carried by the DN ( or C, respecevely) then binds and results in the appropriately- colored signal (red or green, respecevely)

10 Calling Genotypes The raw signal intensiees from the SNP array can be noisy How to cope with the noise Pool the raw signal intensiees from muleple individuals for each SNP and perform a cluster analysis Three clusters for each of the three possible genotypes (,, ) Each dot represents the raw signal intensity for a SNP for each individual

11 CNV Genotyping with rray CGH Genomic DN from two cell populaeons is differeneally labeled (red and green) and hybridized to a microarray Copy numbers are mostly the same across the chromosome between Test and Ref samples ReducEon by a factor of two in copy numbers Log 2 (red/green) =log 2 (red)- log 2 (green)

12 Overview Genotyping technology - SNPs and copy number variaeons Processing the data from genotyping assays - Linkage disequilibrium - Haplotype inference, phasing - Tag SNP seleceon Preliminary analysis of HapMap data

13 Linkage Disequilibrium (LD) LD reflects the relaeonship between alleles at different loci. Ocen, r 2 (correlaeon coefficient) is used as a measure of LD. Locus Locus

14 asic Concepts Parent 1 Parent 2 "" " " a "" "b X "" " " a "" "b a a b b OR a b a b a b a b b etc High LD -> No Recombination (r 2 = 1) SNP1 tags SNP2 Low LD -> Recombination Many possibilities

15 How to Compute r 2 Individuals SNP1 SNP2 SNP r 2 =1.0 SNP1 SNP2 SNP3 r 2 matrix SNP1 SNP2 SNP r 2 =0.0 R 2 =0.0

16 Linkage Disequilibrium in SNP Data r 2 in SNP data from a populaeon of individuals (lack: r 2 =1, white: r 2 =0) genome PopulaEon 2 PopulaEon 2 genome PopulaEon 1 PopulaEon 1

17 Summary SNP/CNV genotyping technology and genotype- calling methods Linkage disequilibrium in the neighboring loci are due to non- random recombinaeon sites across the genome The level of linkage disequilibrium can be quanefied by r 2