Targeted Re-sequencing of Polyploid Wheat Genome

The size of the wheat genome precludes the large-scale analysis of genetic variation by direct genome sequencing. However, a recently developed method of sequence capture offers the possibility of reducing the cost of sequencing by enriching genomic libraries with valuable targets such as coding and regulatory regions. A Nimblegen capture assay targeting 110 Mb of gene-harboring regions of the wheat genome was used to characterize nucleotide sequence variation in >50 accessions of wheat cultivars and landraces. Enriched genomic libraries for next-generation sequencing were prepared by pooling up to 8 barcoded libraries in a single capture reaction followed by 2x100 bp sequencing on the HiSeq2000 instrument. Each sample was re-sequenced to achieve the average target coverage of about 25-50x for reliable variant calling. About 60-73% of Illumina reads could be mapped to wheat chromosome survey sequence data generated by the IWGSC. More than 99% of targets were represented in each sequenced sample. The average SNP density in the analyzed wheat accessions was about 1 SNP per 500 bp. Although the level of genetic diversity positively correlated with the distance of a gene from centromere in all chromosomes, we observed heterogeneity in the levels of genetic differentiation between homoeologous chromosomal regions. The exome re-sequencing data was used to characterize the patterns of SNP variation across the wheat genome and estimate the extent of genetic differentiation among wheat populations. The developed capture assay provides an opportunity to perform cost-effective targeted re-sequencing of gene space for high-resolution dissection of agronomic traits in wheat.