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In molecular breeding of super rice, it is essential to isolate the best quantitative trait loci (QTLs) and genes of leaf shape and explore yield potential using large germplasm collections and genetic populations. In this study, a recombinant inbred line (RIL) population was used, which was derived from a cross between the following parental lines: hybrid rice Chunyou84, that is, japonica maintainer line Chunjiang16B (CJ16); and indica restorer line Chunhui 84 (C84) with remarkable leaf morphological differences. QTLs mapping of leaf shape traits was analyzed at the heading stage under different environmental conditions in Hainan (HN) and Hangzhou (HZ). A major QTL qLL9 for leaf length was detected and its function was studied using a population derived from a single residual heterozygote (RH), which was identified in the original population. qLL9 was delimitated to a 16.17 kb region flanked by molecular markers C-1640 and C-1642, which contained three open reading frames (ORFs). We found that the candidate gene for qLL9 is allelic to DEP1 using quantitative real-time polymerase chain reaction (qRT-PCR), sequence comparison, and the clustered regularly interspaced short palindromic repeat-associated Cas9 nuclease (CRISPR/Cas9) genome editing techniques. To identify the effect of qLL9 on yield, leaf shape and grain traits were measured in near isogenic lines (NILs) NIL- qLL9  CJ16 and NIL- qLL9  C84 , as well as a chromosome segment substitution line (CSSL) CSSL- qLL9  KASA with a Kasalath introgressed segment covering qLL9 in the Wuyunjing (WYJ) 7 backgrounds. Our results showed that the flag leaf lengths of NIL- qLL9  C84 and CSSL- qLL9  KASA were significantly different from those of NIL- qLL9  CJ16 and WYJ 7, respectively. Compared with NIL- qLL9  CJ16 , the spike length, grain size, and thousand-grain weight of NIL- qLL9  C84 were significantly higher, resulting in a significant increase in yield of 15.08%. Exploring and pyramiding beneficial genes resembling qLL9  C84 for super rice breeding could increase both the source (e.g., leaf length and leaf area) and the sink (e.g., yield traits). This study provides a foundation for future investigation of the molecular mechanisms underlying the source⁻sink balance and high-yield potential of rice, benefiting high-yield molecular design breeding for global food security.

Enhanced Expression of QTL qLL9/DEP1 Facilitates the Improvement of Leaf Morphology and Grain Yield in Rice