OBJECTIVETo construct the recombinant adenovirus vector expressing human endostatin.

METHODSHuman endostatin gene extracted from pGEM-T Easy vector containing the target gene fragment was successfully amplified using PCR and cloned into pShuttle2 vector. The target gene was subcloned into an adenovirus vector and the resulted recombinant adenovirus (Ad-hEndo) was linearized before transfected into HEK 293 packaging cells. The Ad-hEndo recombinant adenovirus was efficiently amplified in 293T cells and purified by CsCl density centrifugation, and the titer of the virus was determined.

RESULTSThe amplified hEndostatin cDNA was verified by PCR and sequencing, and the resulted virus titer reached 5.2 x 10(9) pfu/ml.

CONCLUSIONThe recombinant adenovirus containing human endostatin gene has been successfully constructed, which may provide important basis for gene therapy research for angiogenesis-dependent diseases.