Genetic Materials Construction Of Heading Date Genes Ghd7 And Ghd7.1 And Phenotype Scoring

Food security has always been the livelihood of people in China.As one of the staple crops,the stable and increased production of rice guarantees food security.As the human population grows,increasing rice yield has always been one of the major issues that scientists are committed to solving the growing demand for food.Heterosis refers to the superior performance of hybrids relative to their parents.The Utilization of heterosis promoted the yield.Shanyou63,derived from hybridization between Zhenshan97 and Minghui63,showing great heterosis on yield,has made great contributions to our country’s food security.Hence,using Shanyou63 as a model to study heterosis is very representative.Previously,we found that there are lots of loci that contribute to Shanyou63 heterosis,among which Ghd7 and Ghd7.1 have a great contribution to the heterosis of the number of spirelets per panicle.The contribution of Ghd7 and Ghd7.1 to heterosis is currently only supported by population-level data,and there is no direct phenotypic verification of genetic material.Ghd7 and Ghd7.1,controlling rice heading date,plant height,and grain number,which belong to the CCT gene family were cloned.The pair of Ghd7 and Ghd7.1 of the Minghui63 allele is functional haplotypes,while in Zhenshan97,the pair of Ghd7 and Ghd7.1 are non-functional haplotypes.Therefore,we use CRISPR/Cas9 editing technology to create Ghd7 and Ghd7.1 mutants in the background of Minghui63,and then cross with Zhenshan97,obtaining a new type Shanyou63 containing target mutant gene,and finally confirm the impact on Shanyou63heterosis of knockout of the target gene through phenotypic scoring.In order to increase the probability of obtaining edit-positive plants,two targets were designed on the exons of Ghd7 and Ghd7.1,and finally,8 mutants of Ghd7 and 8 mutants of Ghd7.1 in the background of Minghui63 were obtained.Among them,7 mutants of Ghd7 are early termination mutation.,and 8 mutants of Ghd7.1 all destroyed the CCT conserved domain of GHD7.1 protein.To score genetic variations of mutations,phenotypic identification of Ghd7 and Ghd7.1 mutants showed that 1)the heading date of Ghd7 mutants was 2.1-3.4days earlier than that of wild type,while the plant height and the spikelet number per panicle were not different from those of wild type.2)The phenotype of Ghd7.1 mutations significant differences from the wild type in a variety of agronomic traits:the heading date is 4.8-12.1 days earlier,and the plant height is 7.7-18.6 cm shorter than the wild type.The spikelet number per panicle is 21.47-32.80 less,and the grain number per panicle is 10.51-32.83 less than that of the wild type.In addition to the construction of CRISPR materials,the second aim is to construct two near-isogenic lines with Minghui63 allelic Ghd7 and Ghd7.1 through hybridization under the background of Zhenshan97 and then cross with Shanyou63,obtaining another type of Shanyou63 containing two copies of Minghui63 allelic target gene,and finally confirm whether the copy number has effects on Shanyou63 heterosis.The near-isogenic line ZS97（Ghd7-M/M）is based on the Ghd7 near-isogenic line constructed in the early stage of our laboratory.Chip sequencing and backcrossing have been performed to clear the background.At present,it has achieved the BC₂F₄ generation.The near-isogenic line ZS97（Ghd7.1-M/M）is based on the recombinant inbred line of Ghd7.1 and has completed a backcross.In this study,a series of genetic materials of Ghd7 and Ghd7.1 were successfully constructed under the background of Minghui63 and Zhenshan97,which laid the foundation for verifying the relationship between Ghd7 and Ghd7.1 and the number of spikelets per panicle heterosis of Shanyou63.