Identification of gene OsZIPH1 related to rice shattering using Bulk Segregant Analysis

Grain shattering is one of the critical traits in rice, influencing harvesting methods and yield. Seeds from varieties that shatter easily often begin to drop before reaching full maturity. Varieties with difficult-to-thresh or non-threshing characteristics are susceptible to having their branches broken and mixed with straw during mechanical harvesting, leading to relatively severe losses. Therefore, developing rice varieties with a moderate degree of shattering, suitable for production applications, is the primary focus in rice breeding. In this study, the F 2:3 generation population from a cross between the easy-shattering weedy rice variety "H21" and the non-shattering japonica rice variety "Longdao 18" was utilized to investigate shattering resistance through phenotypic identification, Bulk Segregant Analysis (BSA), gene editing, and cytological detection. Gene Os01g0935000, a ZOS1-23 - C2H2 zinc finger protein involved in the regulation of shattering, was preliminarily localized and named OsZIPH1 . Mutant phenotype identification showe that the shattering rate of the mutant is 41.98%, while that of the wild type is 84.33%, the decrease is 50.2% compared to the wild type H21. Cytological analysis revealed that mutant spikelets had incomplete abscission layer structures between the lemma and rachis branches, whereas wild-type plants exhibited clearly defined abscission layer structures. The expression level of this gene in the wild type is 2.63 times that in the knockout mutant. Haplotype analysis has revealed that this gene comprises five haplotypes. HAP5 is unique to Oryza ruf ipogon, while HAP1 is found in nearly all japoni ca rice varieties. HAP2 is predominantly present in 70% of indica weedy rice, and HAP3 is primarily associated with 59.37% of indica improved rice. HAP4 is found in SH and BHA weedy rice, indicating that the shattering gene Os01g 0935000 plays a significant role in the domestication process of rice. These findings enhance our understanding of the biological mechanisms underlying rice shattering and are crucial for developing rice varieties with desirable shattering characteristics. • A gene OsZIPH1 related to rice shattering was identified using bulk segregation analysis. • Cytological evidence abscission layer structures between the lemma and rachis branches was exhibited clearly. • The F 2:3 generation population from a cross between the easy-shattering weedy rice variety "H21" and the non-shattering japonica rice variety "Longdao 18" was utilized.