Characteristics Of Photosynthesis And Sub-cellular Localizations Of Rubisco And Rubisco Activase In Antisense Rca Rice Plants

Rubisco catalyzes the first step of both the photosynthetic carbon assimilation and photorespiratory carbon oxidation, and is a key regulatory enzyme of photosynthesis. The catalytic sites of Rubsico need to be activated by the carbamylation of a lysine residue with the sites follwed by binding of magnesium. The activation of Rubsico in vivo is regulated by Rubisco activase (RCA). The enzyme is a nuclear-encoded chloroplast protein that modifies the conformation of Rubisco, releases inhibitors from the active sites, and increases enzymatic activity. In this study, we regenerated transgenic rice plants with an antisense gene for RCA, and investigated the effects of the reduction of RCA content on photosynthesis, energy dissipation and cellular localization of Rubisco and RCA of rice leaves. The results are as follows:RCA gene (rca) was amplified using specific primers and inserted into pGEM T-easy vector, and then cut with EcoRI after confirming by sequencing. The fragment was subcloned into pBluescript KS+, digested with the enzyme BamHI and inserted into the binary expression vector pCAMBIA1301, and the resulting construction with antisense rca was named pCAMR02. The pCAMR02 vector was introduced into Agrobacterium tumefaciens strain EHA105 by electroporation and transformed to embryos of rice (Oryza. Sativa L.ssp.japonica) cultivar Zhonghuall via Agrobacterium tumefaciens system. Plantlets were regenerated in vitro by resistance selection on medium containing various concentrations of hygromycin. Both GUS histochemical assays and PCR amplification demonstrated that antisense rca was integrated into T₀ genomes and inherited to T₁. The measurement of phenotypes of transgenic rice plants with antisense rca showed that most of them could hardly survive at ambient CO₂ concentration even could not grow. The antisense plants that survived under natural condition were dwarf and grew slower than the wild-type controls.The photosynthetic rate was largely unaffected by RCA concentration until it...