Objective: To construct VEGF gene-targeted small interfering RNA (siRNA) and its expression vector driven by CMV promoter and to investigate its interference effect. Methods: The VEGF gene-targeted hairpin siRNA was designed,two complementary oligonucleotide strands were synthesized.After annealing,two-strand oligonucleotide was inserted into pDC311-SV40-RC vector,which was then identified by PCR and sequenced.Then human U-2 OS cell line was transfected with the vector using lipofectamine method.Finally,ELISA was performed to evaluate the expression of VEGF protein. Results: PCR-identification of positive clone and sequencing confirmed the vector containing the target siRNA.ELISA showed that compared with the control group,the expression levels of VEGF protein in transfected U-2 OS cells were decreased significantly(P＜0.05). Conclusion: VEGF gene-targeted siRNA and its vector mediated by CMV promoter were successfully constructed,which can reduce the VEGF protein expression after transfecting.