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.

Identification of Rice Blast Loss-of-Function Mutant Alleles in the Wheat Genome as a New Strategy for Wheat Blast Resistance Breeding.

Blast is caused by the host-specific lineages of the fungus Magnaporthe oryzae and is the most important destructive disease in major crop plants, including rice and wheat. The first wheat blast outbreak that occurred in Bangladesh in 2016 and the recent epidemic in Zambia were caused by the M. oryzae Triticum (MoT) pathotype, a fungal lineage belonging to M. oryzae. Although a few reported wheat cultivars show modest resistance to MoT, the patterns of genetic variation and diversity of this pathotype make it crucial to identify additional lines of resistant wheat germplasm. Nearly 40 rice blast resistant and susceptible genes have so far been cloned. Here, we used BLAST analysis to locate two rice blast susceptible genes in the wheat reference genome, bsr-d1 and bsr-k1, and identified six identical homologous genes located on subgenomes A, B, and D. We uncovered a total of 171 single nucleotide polymorphisms (SNPs) in an ethyl methanesulfonate (EMS)-induced population, with mutation densities ranging from 1/1107.1 to 1/230.7 kb through Targeting Induced Local Lesions IN Genomes (TILLING) by sequencing. These included 81 SNPs located in exonic and promoter regions, and 13 coding alleles that are predicted to have severe effects on protein function, including two pre-mature mutants that might affect wheat blast resistance. The loss-of-function alleles identified in this study provide insights into new wheat blast resistant lines, which represent a valuable breeding resource.