Targeted Analysis of Polyploid Wheat Genome by Exon Capture

Sequence capture is an effective technique for re-sequencing targets of interest even in complex genomes. Reduction of DNA sample complexity by enriching it with valuable targets allows for analyzing a large number of indiviuals at a relatively low cost. A liquid phase sequence capture was used for obtaining exonic sequences from diploid, tetraploid and hexaploid wheat genomes. Differentially barcoded genomic libraries enriched for coding regions were sequenced using Illumina GAII instrument and generated reads were mapped to exonic reference. The data was used to catalogue inter-genomic divergence, nucleotide sequence polymorphism, gene copy number variation and presence/absence polymorphisms (PAVs) in wheat genome. Our results suggest that EST/cDNA sequences can provide information for designing successful capture experiments for organisms with less developed genomic resources. The impact of polyploidy and intra-genomic gene duplications on the efficiency of variant discovery in the wheat genome was investigated by empirically validating identified variable sites. The depth of read coverage across genes and at variable sites was used to detect gene CNV.  Up to 1% of discovered SNP variants in polyploid wheat were loss-of-function mutations.  Patterns of synonymous and nonsynonymous variation suggested relaxation of selection after origin of polyploid wheat. Inter-genomic divergence data indicated a historical selective constraint in the diploid ancestors of the wheat genomes that acted on genes important to metabolic processes. We demonstrated that, even though the level of sequence similarity between homoeologous genomes and capture baits can bias enrichment efficiency, sequence capture is a powerful approach for targeted variant discovery in wheat exome.