Cloning,Functional Analysis And Application Of Rice RbcS Gene Promoter

The development of biotechnology promotes the transgenic commercialization process. In the meantime, transgenic plants and their products become the central issue on the environment and health. The realization of the directional, safe and efficient expression of foreign gene in plants is one of the key problems to be solved. The cloning of specific promoters expressed in specific organ or developmental stage, the construction of efficient expression vectors, and the controllable genetic transformation are the important ways to solve this problem.The research mainly focus on sequences cloned of ribulose-1,5-bisphosphate carboxylase small Subunit genes ( rbcS ) gene promoter of rice and Arabidopsis thaliana; construction of plant expression vectors; functional analysis of rice rbcS promoter; construction Osrbcs-crylC gene expression vector; identification of transgenic plants. The main results were as follows:1. The promoter sequences of ribulose-1,5-bisphosphate carboxylase small Subunit genes (rbcS ) of rice and Arabidopsis thaliana were cloned by PCR. Compared the tow promoter sequences, we found there were some same light responsive elements in both promoters; also there were some differences between them.2. The cloned Osrbcs and ats1A promoters were fused with gus gene and plant expression vectors were constructed respectively, then introduced into rice by Agrobacterium-mediated transformation. The transgenic plants designated as 1301r and 1301a. The result of GUS histochemical staining showed that gus gene driven by Osrbcs promoter expressed in green tissues, while in 1301a, gus gene expressed in leaf and sheath, but not in culm. Quantitative analysis of GUS activity showed that the gus gene expression level of 1301r was stronger than 1301a.3. 5' end deletions of Osrbcs promoter with a length of 62, 344, 550, 941, 1320bp were fused with gus gene and plant expression vectors were constructed respectively. Quantitative analysis of GUS activity showed that the expression level of gus gene was reduced with the deletion of promoter.4. In our experiment we also found that light induction could enhance GUS activity, but light could not enchance the expression of the 62bp promoter fragment.5. Electrophoretic mobility shift assays showed that I box, GT1 factor, T box and GATA box...