mRNA-Seq for homoeologous gene expression analysis in hexaploid bread wheat

Bread wheat (Triticum aestivum) has a large, complex and hexaploid genome consisting of A, B and D homoeologous chromosome sets. Therefore each wheat gene potentially exists as a trio of A, B and D homoeoalleles, each of which may contribute differentially to wheat phenotypes. We describe a novel approach combining wheat cytogenetic resources (chromosome substitution ‘nullisomic-tetrasomic’ lines) with next generation deep sequencing of gene transcripts (RNA-seq), to directly and accurately identify homoeologue-specific single nucleotide variants and quantify the relative homoeoallelic contribution to gene expression. We discover, based on a sample comprising ~5-10% of the total wheat gene content, that at least 40% of wheat genes are expressed from all three distinct homoeoalleles. Most of these genes show strikingly biased expression patterns in which expression is dominated by a single homoeoallele. The remaining ~60% of wheat genes are expressed from either one or two homoeoalleles only, through a combination of extensive transcriptional silencing and homoeoallele loss. We conclude that wheat is tending towards functional diploidy, through a variety of mechanisms causing single homoeoalleles to become the predominant source of gene transcripts. This discovery has profound consequences for wheat breeding and our understanding of wheat evolution.