The construction of adenovirus vector carrying human tissue inhibitor of metalloproteinase-2 gene

Objective To construct an adenoviral vector carrying human tissue inhibitor of metalloproteinase-2 (TIMP-2) gene for gene therapy. Methods A recombinant adenovirus (AdhTIMP-2) containing human TIMP-2 cDNA fragment was generated by homologous recombination in BJ5183 bacteria. Recombinant plasmids were screened by alteration of antibiotic. The adenovirus vector was then packaged and amplified in 293 cells. The expression of TIMP-2 was detected by the techniques of Western blot and RT-PCR. Results The recombinant adenoviral vector carrying human TIMP-2 was constructed. The titer was 4×10 11pfu/ml after purification. The expression of TIMP-2 gene in 293 cells was detected by RT-PCR. After the 293 cells were transfected with AdhTIMP-2 24 hours, TIMP-2 protein could be detected in the medium by Western blot. Conclusions The recombinant adenoviral vector carrying human TIMP-2 is successfully constructed and paved the way for further application in vascular disease gene therapy.