Grain size is a crucial factor influencing both rice yield and appearance quality. In this study, we identified GL6.1 as a gene associated with rice grain size by map-based cloning. The GL6.1 encodes a protein with CC-NB-LRR domain, a structural motif related to rice disease resistance. Genetic transformation experiments and the construction of gene pyramiding lines indicate that GL6.1 functions as a negative regulatory factor for rice grain length. By the analysis of SSSLs with diverse donor parents, a total of 11 single nucleotide polymorphisms (SNPs) are identified that are closely associated with the phenotype variations. A yeast one-hybrid library screening revealed that GL6.1 interacts with OsWRKY53, a transcription factor associated with rice disease resistance. Furthermore, RNA-seq assay also revealed the involvement of pathways associated with disease resistance, hinting at a multifaceted role for GL6.1 in both yield and stress tolerance traits in rice. These results indicate that the cloning and in-depth studies of GL6.1 would provide valuable insights into the interplay among high yield, superior quality, and stress tolerance, which are critical goals for rice breeding.

Grain size is a key factor influencing grain yield and appearance quality in rice. We identified twelve quantitative trait loci (QTL) for grain length (GL), nine for grain width (GW), and nine for 1000-kernel weight (TKW) using GLU-SSSLs, which are single-segment substitution lines with Oryza glumaepatula as donor parent and Huajingxian 74 (HJX74) as recipient parent. Among the QTL, qGL1-2, qGL1-4, qGL9-2, qGW2-2, qGW9-1 and qTKW9-2 contributed to high grain yield. GL9 was identified as a candidate gene for qGL9-2 by map-based cloning and sequencing, and is a novel allele of GS9. The kernel of NIL-gl9 was slenderer and longer than that of HJX74, and the TKW and grain yield per plant of NIL-gl9 were higher than those of HJX74. The proportion of grain chalkiness of NIL-gl9 was much lower than that of HJX74. Thus, gl9 increased grain yield and appearance quality simultaneously. Three pyramid lines, NIL-gs3/gl9, NIL-GW7/gl9 and NIL-gw8/gl9, were developed and the kernel of each was longer than that of the corresponding recipient parent lines. The gl9 allele may be beneficial for breeding rice varieties with high grain yield and good appearance quality.