Cloning And Functional Analysis Of OsrbcS3 Candidate Candida Candidate Gene At Seedling Stage In Rice

Rice is one of the most important food crops in the world, more than half of the world’s population with rice as the staple food. Heilongjiang Province is located in the north of China,it is the most important production base of grain in China, however, low temperature and cold damage is one of the main obstacles to lead to reduced rice production in Heilongjiang Province. With the expansion of the rice direct sowing area, seedling chilling injury is more and more attention by researchers of rice.Therefore, it plays an important role to increase crop resistance to adversity stress which we study the cold tolerance gene of rice seeding stage and proven resistant to cold molecular mechanism of rice seeding stage.In this experiment, cold tolerance of rice varieties Kongyu131(K131) and cold sensitive varieties Longjing11(L11) as the research objects,from the rice digital expression spectrum, we selected a response to cold tolerance gene Osrbc S3, it is one of the genes involved in photosynthesis.Using the method of PCR,Real-Time PCR, bioinformatics analysis,determination of physiological index,Gateway cloning technology and agrobacterium mediated,studied Osrbc S3 gene at the level of DNA,m RNA and protein. The main results are as follows:1. The significant difference was found in seedling cold resistance of the tested 12 rice varieties of the first, second and third accumulated temperature zone in Heilongjiang province, we screened the cold resistance of rice K131 and cold sensitive L11 used in experiments.2. Through the cold tolerance correlation analysis in bud stage,seedling stage and booting stage in different varieties of rice,we can know seedling stage and bud stage, seedling stage and booting stage of cold tolerance was significant positive correlation.3. Osrbc S3 gene sequences are the same in rice K131 and L11 cloning and NCBI, and the base sequence is exactly the same between K131-Osrbc S3 and L11-Osrbc S3, their CDS area are 528 bp and encoding 175 amino acids.4. 0h as contrast check,we analysis the Osrbc S3 gene expression of cold treatment 2h, 5h, 12 h and 24 h,the results show that Osrbc S3 gene up-regulated expression under cold stress,the expression of K131 Osrbc S3 gene is higher than L11 Osrbc S3 gene,it is probably the reason for the difference between the two varieties of cold resistance.5. The determination of Rubisco activity and other physiological indicators of K131 and L11 under the low temperature, Again Osrbc S3 differences in gene expression quantity is the cause of K131 cold resistance is stronger than L11 been proven again.6. Genetic transformation callus of L11,obtain transgenic plants, preparation of materials for the verification Osrbc S3 gene.