A MADS51 allele is associated with flowering-time divergence and contributes to ecological adaptation during wild rice speciation.

Flowering-time divergence can contribute to ecological speciation, but causal genes and mutations remain largely unknown in wild plants. Here, we investigate flowering-time divergence between the wild rice species Oryza rufipogon and O. nivara. By establishing a transformation system for wild rice and combining genome scans, population genomics, and functional assays, we identify MADS51 as a contributor to flowering-time divergence. Knockout of MADS51 in O. rufipogon accelerates flowering, whereas overexpression delays flowering. A functional haplotype (functional haplotype 2 [FH2]), defined by three nonsynonymous substitutions and a 4-bp frameshift, is nearly fixed in Southeast Asian O. nivara. Complementation assays confirm that FH2 promotes earlier flowering than ancestral haplotypes. Population genetic analyses reveal a selective sweep associated with FH2, which appears to have arisen de novo in O. nivara. These findings provide insights into how de novo mutation can contribute to phenotypic differentiation in wild rice.