Candidate gene FLBOA9 identified by genome-wide association acts as a negative regulator for leaf angle of functional leaves in rice

With the continuous growth of the global population, steadily increasing rice yield is crucial for ensuring food security. The leaf angles of the top three functional leaves of rice directly determine the canopy structure and photosynthetic capacity of the population. Therefore, optimizing the angles of functional leaves is a key approach to developing an ideal plant type and boosting rice yield. Nevertheless, the complex genetic basis governing this trait remains to be further elucidated. In this study, 437 rice accessions were selected to detect the candidate genes controlling the base and opening angle of the top three functional leaves by genome-wide association study (GWAS) based on 3,172,711 high-quality single nucleotide polymorphisms (SNPs). A total of 44 significant loci were identified in both the general linear model (GLM) and the mixed linear model (MLM). Among them, 36 loci were newly discovered in this study. At the same time, two significant sites were co-located in the base and opening angle of the third leaf from the top, which were renamed as qFLBOA3 and qFLBOA9 . Based on functional annotation and haplotype analysis of the genes in the physical location of QTLs, the gene Functional Leaf Base and Opening Angle 9 ( FLBOA9 ) was verified by CRISPR/Cas9 technology as a negative regulatory factor of the leaf angle trait in functional leaves of rice. On this basis, multi-gene pyramiding breeding prediction was further performed using the elite alleles of FLBOA9 , LG5 , LC2 , and FIB under three crossing strategies, which theoretically project further reductions in leaf angle. This study demonstrates that FLBOA9 is a key negative regulator of functional-leaf angle, and based on the results of superior alleles, three hybridization schemes were designed to conduct multi-gene pyramiding prediction, aiming at developing compact plant architecture for improving light‑use efficiency and further raising grain yield.