Molecular Cloning And Expression Patterns Exploration Of OsWRKYS2Gene Encoding Starch Synthesis

Rice quality improvement has become an important part of rice breeding，Amylose content isone of the most important characters to improve rice quality, It was confirmed that the gene ofsynthesis starch is induced by Sugar signal. The representative gene named SUSIBA2can combinewith promoter related to amylopectin synthesis of starch synthase as a transcriptional activator.However, according to GenBank sequence that has been speculated and using cDNA of the rice（Nipponbare）as the template, We have successfully obtained WRKY transcript gene namedOSWRKYS2using RT-PCR method. Eventually,In order to clarify molecular regulatory mechanismof rice starch synthesis, the molecular cloning and biological function of OSWRKYS2gene wereanalysed, and made accumulation and composition of starch be changed, Consequently, thepossibility of factory-like prodution to improve rice quality using genetic engineering is to realize.Here are results as follow:1. Based on comparative analysis database in indica-japonica rice, The WRKY transcriptgene OSWRKYS2encoding rice starch synthesis was cloned. The cDNA with the length of1731bpencoded570aminoacids(aa), and was included a typical WRKYGQK conservative structuredomain and a zinc finger structure, and belongs to the WRKY family group I, Then bioinformaticssoftware predicted that there may be no signal peptide, and located in the mitochondrial matrix..2. To confirm the function of OsWRKYS2gene, The P3300plasmid was exactly digested byrestriction enzyme (BamHI/SacI) and sequence reaction, and combined with OsWRKYS2gene byhigh fidelity enzymes. The recombinant plasmid named p3300-Ubi-OSWRKYS2were constructedsuccessfully as expectation.3. OsWRKYS2gene was transformed into rice by the agrobacterium-mediated method,184strainswere transformed into the rice, and only28strains resistant plants were obtained, The conversionefficiency was8%.And harvest the T0generation of genetically modified seeds.UsingPCR-Southern Analysis, we confirmed that OsWRKYS2gene was expressed in rice with the singlecopy number, and28strains positive plants were received. 4. Molecular surveillance confirmed that amylase content increased4.96%than the wild type,and kasalath increased by4.73%, Amylopectin content than wild type have increased significantly.The increased expression of starch branching enzyme (SBE),soluble starch synthase(SS),and AD-glucosepyro-phosphorylase(AGPP) activity were higher than that of wild type, and the three kindsof enzymes in the grain starch synthesis plays an important role in the process, Presumably thegenes can start starch synthesis gene expression in the sugar signal transduction pathways, andwhich affects the expression of key enzyme activity in starch synthesis, but the rice quality ofpractical application also needs to be further explored...