Phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) is located at the key site of plant carbon metabolism pathways which can regulate the protein content and the lipid content in cell, and the down-regulation of PEPC expression caused increased lipid accumulation. In this study, we cloned the fragment of the "bacterial-type" pepc2 gene from Chlamydomonas reinhardtii (named Crpepc2)and the Hsp70A-RBCS2 promoter (named HR), inserted the HR and Crpepc2 into pSP124s vector, then the reverse recombinant vector pSP124s-HR-reve-Crpepc2 was obtained. The pSP124s and pSP124s-HR-reve-Crpepc2 vector was transformed into C. reinhardtii cc-503 strain through biolistic, respectively, the blank strain and reverse stain was obtained. The relative expression of pepc2 gene was measured in wild stain, blank strain and reverse stain by using qPCR. The date showed that the pSP24s in the blank strain didn’t obviously influence the relative expression of pepc2 gene, which was 92.95% of that of the wild stain, and the reve-Crpepc2 in the reverse stain significant inhibition the relative expression of pepc2 gene, which was only 2.94% of that of the wild stain. This result indicated that we have established the method to detect the relative expression of pepc2 gene in C.reinhardtii by using qPCR, and it also proved that “reverse vector method (RVM)” could effectively inhibit the expression of pepc2 gene in C.reinhardtii. These efforts were lay a good foundation for screening high-lipid content reverse strain.