Rice is an important food crop, providing staple food for more than half of the world’s population. Panicle traits are the main factors affecting rice yield. Discover the elite haplotype of the panicle regulation gene, and provide important germplasm and gene resources for pyramiding breeding.

In this study, recombinant inbred lines (RILs) derived from a cross between SN265 and R99 were re-sequenced through high-throughput sequencing. QTL analysis and candidate gene identification were conducted on the grain number on the primary branch, the grain number on the secondary branch, and the grain shape. The sequences of candidate genes were compared using the long-read sequence assemblies of SN265 and R99. The combination of candidate genes that can maximize grain yield was selected among RILs. Finally, the super rice variety SN265 was improved using CRISPR/Cas9 gene editing technology.

The R99 had significantly more grain number per panicle and grain number on the secondary branch, whereas SN265 had significantly more grain number on the primary branch. The grain of R99 is slender, and the grain of SN265 is short and round. The RILs were sequenced with approximately 6.25-fold depth. For parent lines, 30.0-fold depth and 32.0-fold depth data were generated for R99 and SN265, respectively. Subsequently, a bin map was constructed by 1456445 high-quality SNPs. The genetic map containing 3 569 recombinant blocks, with an average length of 58.17 kb. The QTL analysis detected a QTL on Chr.9 for grain number per panicle and grain number on both primary and secondary branch, a QTL on Chr.1 for grain number per panicle and grain number on the secondary branch, a QTL on Chr.5 for grain shape. The candidate gene prediction and sequence comparison showed that DEP1 regulated the grain number on both primary and secondary branches of rice, Gn1a mainly regulated the grain number on secondary branches of rice, and qSW5 mainly regulated the grain shape. The yield of the combination of Gn1a^R99/DEP1^SN265/qSW5^SN265 alleles showed an advantage in yield performance among the RILs. We further conducted a molecular design breeding to SN265 by knocking out the Gn1a locus using CRISPR/Ca9 gene editing technology, and the grain number per panicle of the transgenic plants increased significantly compared to that of SN265.

This study used RILs derived from a XI/GJ cross and high-throughput sequencing technology to conduct QTL analysis of rice panicle traits, revealed the effects of DEP1, Gn1a, and qSW5 on grain number per panicle and grain shape, and clarified that Gn1a^SN265/ DEP1^R99/qSW5^R99 was the best gene combination in RILs. The yield per plant was further improved by knocking out the Gn1a locus of SN265. This study provided important germplasm and gene resources for pyramiding breeding with elite alleles.