GWAS Combined with Meta-Analysis Identifies OsDREB6 as a Regulator of Coleoptile Elongation Under Anaerobic Conditions in Rice.

Identifying genes resistant to anaerobic germination can provides key genetic targets for breeding direct seeding rice varieties with anaerobic tolerance. In this study, genome-wide association analysis (GWAS) was performed on coleoptile length (CL) of 591 natural rice populations under anaerobic conditions, and a total of 34 significant QTLs were identified, with eight of them co-localized with previous studies. Furthermore, through meta-analysis of 156 initial QTLs from 21 independent studies related to anaerobic germination, 37 MQTLs were identified, including 4 core MQTLs. Integration of GWAS with meta-analysis revealed the overlap between the physical interval of qCL9.5 on chromosome 9 and MQTL9.2, highlighting it as a reliable locus. Notably, our analysis pinpointed the dehydration-responsive element-binding protein 6 gene, OsDREB6, as a potential regulator impacting anaerobic germination in rice seeds. Phenotypic analysis revealed that the ko-osdreb6-1 and ko-osdreb6-2 mutants exhibited significantly increased CL and germination sprout length under aerobic treatment for 4 days compared to WT. In contrast, disruption of OsDREB6 caused reduced CL in plants seeds under under anaerobic 4-day treatment and anaerobic 3-day treatment after seed dehiscence. Additionally, the relative coleoptile lengths of the mutants after 4 days between anaerobic and aerobic treatments were significantly lower than those of WT. RNA-seq and MapMan analysis of the ko-osdreb6-1 suggested that OsDREB6 may regulate the coleoptile elongation under anaerobic conditions by affecting the expression of related genes involved in the sucrose and starch metabolism. Overall, our study demonstrated that the effectiveness of combining GWAS with meta-analysis of QTL in identifying genetic loci and key genes for improving anaerobic germination tolerance in direct seeding rice breeding.