Cloning And Expression Analysis Of CPR5 Gene And Promoter In Rice

The plant is frequently subject to environmental stress in the growth, and heat and salt stress is one of the primary factors. These stresses can limit plant growth, and are often responsible for low yield and poor quality of plant. The results from the previous study indicated that AtCPR5 is not only disease-resistant but also heat-resistant. In the paper, through sequence comparison of AtCPR5 which is a pleiotropy gene with rice geneome ,we obtained a highly similar gene,named OsCPR5. We fisrtly predicted the potential functions of OsCPR5 gene by bioinformation, amplified OsCPR5 gene and its promoter and constructed expression vectors,then studied their functions in Arabidopsis , which purpose is to understand whether OsCPR5 is heat-resistant or not and the stress responses mechanism in rice .The main results were as follows:(1)Through sequence analysis, OsCPR5 may be a resistant gene to stress responses in rice: Sequence analysis of amino acid demonstrated both OsCPR5 and AtCPR5 have five transmembrane domains and high similarity of protein secondary structure。Further analysis of OsCPR5 promoter Cis-acting element demonstrated that OsCPR5 gene promoter contains a variety of stress-responsive elements, such as HSE, MBS, TC-rich repeats, ABRE,EIRE and W box, etc. These data suggest that OsCPR5 is not only a pleiotropy gene,but also plays an important role in response to various stresses in rice.(2) Recombination plasmid pBI-OsCPR5P-GUS and pCanG-OsCPR5 were obtained and transformed into EHA105: Total DNA was isolated from 2-week-old rice seedling and the sequence of OsCPR5 promoter was amplified by PCR, and then was inserted pBI101 vector which contains reporter gene of GUS. OsCPR5(Os01g68970)cDNA was amplified and inserted the binary vector of pCanG which contains 35S promoter of cauliflower mosaic virus. Finally both recombination plasmid named pBI-OsCPR5P-GUS and pCanG-OsCPR5 were transformed into EHA105, respectively.(3)OsCPR5P-GUS and 35S-OsCPR5 transgenic plants were obtained: Two recombination plasmids were introduced into wild-type of Arabidopsis by Agrobacterium tumefactions using floral dip method. OsCPR5P-GUS and 35S-OsCPR5 transgenic plants were obtained by kanamycin screening and PCR analysis.(4) Gus staining of OsCPR5P-GUS transgenic plants indicated that OsCPR5 promoter express only in cotyledon and euphylla and is a tissue-specific promoter. Simutaneously, the expression patterns of OsCPR5 is very similar to AtCPR5.