Genome-wide association analysis identifies candidate miRNA-targeted genes associated with spatial grain size and...

Grain homogeneity in size and weight is crucial for improving grain quality. Reducing spatial variability in grain size and weight across the spike is important for increasing grain uniformity, quality, and breeding barley with consistent yield. However, there is limited knowledge on the molecular genetic basis of grain shape in temperate cereals, including barley. Therefore, we studied grain size and weight along the barley spikes, comparing apical vs. basal grains in both two- and six-rowed spikes and central vs. lateral grains in six-rowed spikes. Regardless of row type, the basal part of the barley spike has more uniform grains. The grains from two-rowed barley had significantly wider and higher thousand-kernel weight, while grains from six-rowed barley were longer with increased grain area. The results showed a clear basal-to-apical gradient in grain size and weight, which implies genetic control of the variation. Gene-based analysis identifies a family of miRNA-regulated transcription factors, SQUAMOSA PROMOTER BINDING PROTEIN-LIKEs (HvSPLs) and APETALA2/Ethylene Responsive Factor (HvAP2/ERF), that are associated with grain size-related traits in barley, establishing a novel connection between positional grain characteristics and miRNA-driven regulation mechanisms in a temperate cereal. The results provide an important foundation for uncovering the biological functions of HvSPLs and HvAP2/ERF, which hold important potential for regulating agronomic traits and for understanding their roles in temperate cereals like barley. Genetic variation in miRNA-targeted HvSPL and HvAP2/ERF genes will provide more natural trait-enhancing alleles for crop breeding.