A large‐scale whole‐exome sequencing mutant resource for functional genomics in wheat

Hexaploid wheat (Triticum aestivum), a major staple crop, has a remarkably large genome of ~14.4 Gb (containing 106 913 high‐confidence [HC] and 159 840 low‐confidence [LC] genes in the Chinese Spring v2.1 reference genome), which poses a major challenge for functional genomics studies. To overcome this hurdle, we performed whole‐exome sequencing to generate a nearly saturated wheat mutant database containing 18 025 209 mutations induced by ethyl methanesulfonate (EMS), carbon (C)‐ion beams, or γ‐ray mutagenesis. This database contains an average of 47.1 mutations per kb in each gene‐coding sequence: the potential functional mutations were predicted to cover 96.7% of HC genes and 70.5% of LC genes. Comparative analysis of mutations induced by EMS, γ‐rays, or C‐ion beam irradiation revealed that γ‐ray and C‐ion beam mutagenesis induced a more diverse array of variations than EMS, including large‐fragment deletions, small insertions/deletions, and various non‐synonymous single nucleotide polymorphisms. As a test case, we combined mutation analysis with phenotypic screening and rapidly mapped the candidate gene responsible for the phenotype of a yellow‐green leaf mutant to a 2.8‐Mb chromosomal region. Furthermore, a proof‐of‐concept reverse genetics study revealed that mutations in gibberellic acid biosynthesis and signalling genes could be associated with negative impacts on plant height. Finally, we built a publically available database of these mutations with the corresponding germplasm (seed stock) repository to facilitate advanced functional genomics studies in wheat for the broad plant research community.