SNP discovery and evaluation with whole genome re-sequencing using pooled DNA in Japanese Black cattle

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  • 黒毛和種のDNAプールを用いた全ゲノムリシーケンシングによる網羅的SNP探索とその評価
  • クロゲワシュ ノ DNA プール オ モチイタ ゼン ゲノムリシーケンシング ニ ヨル モウラテキ SNP タンサク ト ソノ ヒョウカ

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Abstract

It has become evident that SNPs are useful DNA marker in bovine genome research, such as genome-wide association study and family-based linkage analysis. Next step will be to construct bovine SNP database containing million of common SNPs (minor allele frequency, MAF 0.05). The database will facilitate to fine-map regions of interest and to exclude common SNPs for discovery of rare, deleterious variants. Massively parallel sequencing enabled to discover SNPs comprehensively from whole genome or targeted genome region(s) with reduced cost and period. Although, million of SNPs are detected, it is cost-prohibitive to estimate their allele frequencies followed by Sanger sequencing. However, re-sequencing using pooled DNA enable to estimate MAF, based on read depth for each allele. We performed SNP discovery by deep whole genome re-sequencing. Fifty-two steers were selected as a Japanese Black cattle representative population, based on principal components analysis using the BovineSNP50 (Illumina) genotypes, and classified into 4 lines according to the pedigree information. Genomic DNA was pooled in each line and sequenced with 2 x 40-bases reads using Genome Analyzer II (Illumina). The data obtained from 15 runs in 4 lines (159.4 Gb ; 59.70 x) were analyzed using UMD3.0 as a reference sequence, and 13.3 million SNPs (estimated MAF 0.05) with a minor allele supported by 2 reads were detected on the autosomes. Four million six hundred thousand of these SNPs are located in non-repetitive regions, suggesting a unique SNP (average interval ; 596-bp). The estimated MAF by pooled DNA showed a strong correlation with MAF from Bovine HD Chip (Illumina) genotype. We expect that most of common SNPs in Wagyu will be detected with estimated MAF in each line as well as in a general population, which will provide useful information for a genomic analysis of traits.

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