The Construction Of High Efficient Expression Vector And Its Expression In Chlamydomonas Reinhardtii Chloroplast

Chlamydomonas reinhardtii had a special cell structure, clearly geneticbackground and it ofen was used as a model organism for the study of life pHenomena.C. reinhardtii chloroplast was regarded as a good bioreactor and ofen used to expressvaluable proteins. But the low expression levels of foreign genes hindered thecommercial application of Chlamydomonas reinhardtii.The promoters and enhancers were the key elements to regulate the expression offoreign genes. The activity of the promoters and enhancers could affect the expressionlevels of foreign proteins. In this paper, in order to improve the expression levels offoreign genes, vectors using the Green Fluorescent Protein (gfp) as reporter gene andusing promoter atpA or psbA and different enhancers as regulation elements wereconstructed. One of the vector was transformated into the Chlamydomonas reinhardtiiby the method of gene gun (PDS-1000/He).The major contents and results were as following:1、Using the gfp as reporter gene and promoter atpA and enhancers as regulationelements, the C. reinhardtii chloroplast expression vectors were constructed.2、The promoter psbA was cloned using the method of PCR. Then the psbApromoter and different enhancers were used to regulate the express of the reportergene.3、The plasmid pgfpa2containing the genes of gfp and aadA was constructed.Then the plasmid of pgfpa2was transferred into C. reinhardtii using the method ofparticle bombardment. After selected on Spc (spectinomycin) plate, the recombinanttransformant was obtained.The results above established a solid foundation for further work.