CRISPR/Cas9-mediated FeS-ELF3 inactivation overcomes heteromorphic self-incompatibility in common buckwheat

Common buckwheat (Fagopyrum esculentum) is a climate-resilient pseudocereal; however, its global adoption is constrained by the heteromorphic self-incompatibility associated with distyly. While the S-locus early flowering 3 (FeS-ELF3) gene has been identified as a key regulator of self-compatibility, a stable genetic transformation of F. esculentum has not yet been developed. In this study, we established an Agrobacterium-mediated transformation protocol for F. esculentum by adapting an F. tataricum procedure, incorporating a heat-shock treatment at 43 °C for 5 min. This achieved a transgenic frequency of 27.27% among regenerated plants (6/22) and an absolute editing efficiency of 4.55% (1/22) in the agronomically relevant 'Panda' cultivar. Targeted knockout of FeS-ELF3 via CRISPR/Cas9 yielded a self-compatible line with long-homostylous flowers. Plants derived from this single edited founder event exhibited architectural shifts - specifically reduced height and shorter inflorescences - without an apparent reduction in seed yield under greenhouse conditions. Because these observations stem from a single edited line, further evaluation of independent mutant lines or complementation assays is required to definitively distinguish FeS-ELF3-specific pleiotropic effects. Nevertheless, this study provides a proof-of-concept for overcoming self-incompatibility and establishes a genomic framework for the rapid improvement of Fagopyrum esculentum.