Bulked Segregant Analysis Coupled With Whole-Genome Sequencing (BSA-Seq) and Identification of a Novel Locus, qGL3.5, That Regulates Grain Length

Abstract Rice grain length (GL) directly affects the yield and quality of this species. Very few GL-related genes cloned are applied in production because their yield-increasing effect was not obvious. In this study, the two bulk-DNA pools (L-pool and S-pool) and their parents’ (KJ01 and Huaye 4) DNAs were subjected to high-throughput sequencing. After assessing the quality of the data, we obtained a total of 100.22 Gb of high-quality data; the average coverage depth was 55x, and the genome coverage was 96.51%. After combining the association results of the ED and SNP index methods, we mapped the GL genes to a 0.34 Mb “hotspot” region on chromosome 3, which contains 37 genes related to various traits. The 37 predicted genes were further analyzed by the use of the Gene GO, COG database and so on. Thirty-three genes were annotated by GO functions. According to the GO annotations, three genes whose molecular function involved in the plasma membrane and intracellular membrane-bound organelles were detected via CRISPR/ Cas9 editing technology. ORF33 was verified to regulate GL and was the target gene qGL3.5. These results provides a new gene resource for rice grain shape breeding and a starting point for functional characterization of the wild rice GL gene.