Enhancing Rice Grain Quality Through the Knock-out of the OsSPL16 Gene

The OsSPL16 (SQUAMOSA promoter-binding protein like 16) or GW8 (Grain Width 8) gene is expressed specifically in developing seeds and positively controls grain width and weight by modulating cell proliferation and enlargement in the spikelet hull of rice. OsSPL16 acts as the direct upstream suppressor of GW7 (Grain Width 7), a gene responsible for grain slenderness. The down-regulation of the OsSPL16 gene will effectively disrupt its interaction with the GW7 gene, leading to the development of slender rice grains. In our investigation, we introduced grain slenderness to a bold grain-type rice cultivar, ASD16, by employing the CRISPR/Cas9 system to knock-out the function of the OsSPL16 gene. Molecular analysis of spl16 mutants unveiled the existence of mutations within the first exon of OsSPL16, which included in-frame and frame-shift mutations. Our results demonstrated that inducing frame-shift mutations in the coding sequence of the OsSPL16 gene could effectively impart grain slenderness in bold grain varieties.