Exome sequencing of bulked segregants identified a novel TaMKK3-A allele linked to the wheat ERA8 ABA-hypersensitive germination phenotype.

KEY MESSAGE: Using bulked segregant analysis of exome sequence, we fine-mapped the ABA-hypersensitive mutant ERA8 in a wheat backcross population to the TaMKK3-A locus of chromosome 4A. Preharvest sprouting (PHS) is the germination of mature grain on the mother plant when it rains before harvest. The ENHANCED RESPONSE TO ABA8 (ERA8) mutant increases seed dormancy and, consequently, PHS tolerance in soft white wheat 'Zak.' ERA8 was mapped to chromosome 4A in a Zak/'ZakERA8' backcross population using bulked segregant analysis of exome sequenced DNA (BSA-exome-seq). ERA8 was fine-mapped relative to mutagen-induced SNPs to a 4.6 Mb region containing 70 genes. In the backcross population, the ERA8 ABA-hypersensitive phenotype was strongly linked to a missense mutation in TaMKK3-A-G1093A (LOD 16.5), a gene associated with natural PHS tolerance in barley and wheat. The map position of ERA8 was confirmed in an 'Otis'/ZakERA8 but not in a 'Louise'/ZakERA8 mapping population. This is likely because Otis carries the same natural PHS susceptible MKK3-A-A660S allele as Zak, whereas Louise carries the PHS-tolerant MKK3-A-C660R allele. Thus, the variation for grain dormancy and PHS tolerance in the Louise/ZakERA8 population likely resulted from segregation of other loci rather than segregation for PHS tolerance at the MKK3 locus. This inadvertent complementation test suggests that the MKK3-A-G1093A mutation causes the ERA8 phenotype. Moreover, MKK3 was a known ABA signaling gene in the 70-gene 4.6 Mb ERA8 interval. None of these 70 genes showed the differential regulation in wild-type Zak versus ERA8 expected of a promoter mutation. Thus, the working model is that the ERA8 phenotype results from the MKK3-A-G1093A mutation.

Genome-Wide Association Mapping for Tolerance to Preharvest Sprouting and Low Falling Numbers in Wheat.

Preharvest sprouting (PHS), the germination of grain on the mother plant under cool and wet conditions, is a recurring problem for wheat farmers worldwide. α-amylase enzyme produced during PHS degrades starch resulting in baked good with poor end-use quality. The Hagberg-Perten Falling Number (FN) test is used to measure this problem in the wheat industry, and determines how much a farmer's wheat is discounted for PHS damage. PHS tolerance is associated with higher grain dormancy. Thus, breeding programs use germination-based assays such as the spike-wetting test to measure PHS susceptibility. Association mapping identified loci associated with PHS tolerance in U.S. Pacific Northwest germplasm based both on FN and on spike-wetting test data. The study was performed using a panel of 469 white winter wheat cultivars and elite breeding lines grown in six Washington state environments, and genotyped for 15,229 polymorphic markers using the 90k SNP Illumina iSelect array. Marker-trait associations were identified using the FarmCPU R package. Principal component analysis was directly and a kinship matrix was indirectly used to account for population structure. Nine loci were associated with FN and 34 loci associated with PHS based on sprouting scores. None of the QFN.wsu loci were detected in multiple environments, whereas six of the 34 QPHS.wsu loci were detected in two of the five environments. There was no overlap between the QTN detected based on FN and PHS, and there was little correlation between the two traits. However, both traits appear to be PHS-related since 19 of the 34 QPHS.wsu loci and four of the nine QFN.wsu loci co-localized with previously published dormancy and PHS QTL. Identification of these loci will lead to a better understanding of the genetic architecture of PHS and will help with the future development of genomic selection models.

Registration of Zak ERA8 Soft White Spring Wheat Germplasm with Enhanced Response to ABA and Increased Seed Dormancy.

Zak ERA8 (ENHANCED RESPONSE to ABA8) (Reg. No. GP-966, PI 669443) is a unique line derived from soft white spring wheat (Triticum aestivum L.) cultivar Zak that has increased seed dormancy but after-ripens within 10 to 16 wk. The goal in developing this germplasm was to use increased seed dormancy to improve tolerance to preharvest sprouting, a problem that can cause severe economic losses. This germplasm was developed by USDA-ARS, Pullman, WA, in collaboration with Washington State University. Zak ERA8was tested under experimental number 60.1.27.10. The ERA8mutation was generated by chemical mutagenesis followed by selection for the inability to germinate on abscisic acid (ABA) concentrations too low to inhibit wild-type Zak seed germination. The semidominant Zak ERA8 line has been backcrossed twice to wild-type Zak. Following the first backcross, Zak ERA8 showed similar morphological and grain quality traits to the original Zak cultivar.