| Rice heading stage directly affects photosynthetic efficiency and regional adaptability,which in turn affects rice yield and quality.The regulation of heading stage is conducive to the planting area of excellent germplasm and the exchange of dominant germplasm resources.With the advantage of gene editing technologies,it is of great theoretical significance and application value to study the phenotypic effects of each point in the regulation pathway of heading stage systematically,and to discover new genes for controlling rice heading stage,and to create excellent rice germplasm.1.Establishment of mutant library of genes related to heading stage and investigation of heading difference in riceIn this study,Xiushui 134 was used as material to edit 30 genes related to heading stage in rice,obtaining the mutant materials of each gene to create a mutant library of the entire regulatory pathway.The heading changes of each gene mutations under long-day and short-day were systematically investigated.Hence,the alleles which could be beneficial to northern planting of southern rice varieties were preliminarily screened out.The contribution of each mutant to heading delay from large to small was,ehd2(+nd),sdg723(+36d),osmads50(+30d),oself3(+29d),ehd3(+21d),ehdl(+18d),osfdl(+16d),hd3a(+14d),ehd4(+7d),osfkfl(+3d).While,the contribution to promote the flowering time from large to small was,ghd7(-20d),ghd8(-13d),hd16(-6d).It was suggusted that the mutations of these genes were the expected targets for the practice of rice breeding in latitude-across introduction.2.Preliminary study on the function of OsbZIP74 in the heading stageW2897 was a flowering-late mutant,which was late heading for 28 days and 17 days under natural short-day conditions in Hainan and under long-day conditions in Fuzhou respectively.We constructed the F2 population with the material and 931 1.The mutagenesis gene osbzip74 in the late heading mutant 2897 was cloned by BSA mapping.It was found that there were 44 bases missing in the transmembrane structure of the gene compared with the wild type,and the complementary experimental heading phenotype was recovered.The gene editing materials obtained at the mutation position delayed the rice heading.Further expression level detection revealed that osbzip74 gene was highly expressed in leaves and had daily rhythm characteristics.Under long-day and short-day conditions,it mainly affected the expression of Hd3a gene directly,rather than affected the expression of other upstream genes.Subcellular localization indicated that the nuclear and the cell membrane localization of osbzip74 became a nuclear localization after mutation.The transcriptome results indicated that the locational changes of the mutant gene affected the expression of a series of genes with transmembrane transport-related genes on the membrane,which was consistent with the evidence of the locational changes of the gene.These results suggested that the osbzip74 was a photoperiod gene that potentially affected heading due to the localization changes.3.Creation of elite rice varieties by genome editing technologiesRice breeding in China has gone from yield breeding to quality breeding currently.In this study,we focused on the important quality traits of rice color,aroma and taste.and improved the target phenotypes through genome editing technologies.The white rice was improved to high-quality red rice with significantly increased anthocyanin content.Then the proanthocyanidins and anthocyanins of the improved materials were increased by 5 and 18 times compared with the wild type,respectively.The 2AP concentration of the obtained fragrant rice reached 0.12ng/mg,which was about 18 times that of the wild type,even more than 3 times higher than the positive control NX32(0.0366ng/mg).Finally,we also obtained the new glutinous rice materials with amylopectin contented up to the national standard through CRISPR/Cas9 system. |
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