Using CRISPR/Cas9 Gene Editing Technique To Regulate The Rice G Protein γ Subunit To Improve Rice Yield And Quality

Heterotrimer G protein signaling mediates intracellular responses to external stimuli and is evolutionarily conserved in many organisms.In rice,G protein is involved in the regulation of several important agronomic traits.In this study,two atypical G proteinγsubunits,DEP1 and GS3,were found to antagonize rice yield and quality.The dep1 mutant significantly increased the number of grains per panicle,but adversely affected taste value,cooking and chalkiness related traits.The number of grains per panicle of gs3 mutant decreased,but the grain length increased significantly.Interestingly,gs3 mutant plants performed significantly better than wild-type and DEP1 mutants on food taste,appearance,cooking quality and rapid viscosity analyzer（RVA）.In order to combine the advantages of GS3 and DEP1 and improve the yield and quality of rice,site-directed gene editing was carried out in the GS3 locus of super rice variety Shennong（SN265）.The DEP1 site of SN265 was mutated,with upright panicle and excellent yield,but the quality of food taste was average.Gene editing plant SN265/GS3 increased grain length and further increased yield.Compared with SN265,the texture and RVAs of SN265 were significantly improved,and the taste quality was improved.This study provides a strategy to simultaneously improve rice yield and quality by gene editing G proteinγsubunit.The specific analysis results are as follows:1.Two vectors CRISPR/Cas9-GS3 and CRISPR/Cas9-DEP1 were successfully constructed and transferred into the receptor material Sasanishiki,and transgenic plants with target gene editing were successfully obtained.2.Analysis of the agronomic traits of T₂ transgenic mutant lines showed that under Sasanishiki genetic background,the plant height,grain number per panicle and 1000-grain weight of DEP1 mutant were significantly decreased,while the difference in panicle number and seed setting rate was not significant.The grain length,1000-grain weight and grain number per panicle of GS3 mutants were significantly increased,while the grain number per panicle and seed setting rate were significantly decreased,indicating that the grain number per panicle of DEP1 mutants was significantly increased due to the increase of grain number per panicle.Although the grain length of GS3 mutants was significantly increased,the grain number per panicle was reduced so that the stage yield was similar to that of wild type.3.The glumes cell numbers of Sasanishiki and gs3 mutants were significantly different,and the cell length of GS3 mutants was significantly increased.Chalkiness of Sasanishiki and GS3 mutants was significantly lower than that of DEP1 mutants.Further observation of the morphology and arrangement of endosperm starch grains showed that starch grains of Sasanishiki and GS3 mutants were compact and polygonal.4.Quality analysis of T₂ transgenic mutants showed that gs3 mutants had obvious advantages in appearance,cooking and eating quality except for a slight decrease in milling quality,while DEP1 mutants had similar quality traits to wild type except for a decrease in eating and chalkiness traits.5.In order to improve the quality traits of super rice variety SN265,site-directed gene editing of GS3 locus was carried out.The results showed that there was no significant difference in plant type,plant height,panicle number and seed setting rate between GS3 mutant and SN265,and grain length was significantly increased.Therefore,although the number of grains per panicle decreased,but increased 1000 grain weight,ultimately increased the yield per plant.Although the milling quality of mutants was somewhat decreased,the appearance,cooking and eating taste of mutants were significantly improved.There was no significant difference between the content of direct starch and fatty acid,but the content of protein was significantly reduced.In conclusion,the gs3mutant could significantly improve the quality traits of SN265.