[Construction and identification of matrix metalloproteinase inhibitor-1 siRNA eukaryotic expression vectors].

OBJECTIVE: To construct TIMP-1 siRNA eukaryotic expression vectors and evaluate their effect on TIMP-1 mRNA expression in hepatic stellate cells. METHODS: The combinant lone DNA with cutting sites of BamH I and Xho I enzyme according to the sequences of 447-465, 552-540 TIMP-1 of rats and nonspecific sequence were selected and cloned to pGEM-T vector and sub-cloned to pRNAT-U6.2. They were then identified by double enzyme digestion analysis and DNA sequencing. Three plasmids were transfected into T6 separately through an oligofectamine package. TIMP-1 mRNA expression was evaluated by RT-PCR. RESULTS: Targeting sequences of TIMP-1 siRNA eukaryotic expression vectors were correct. TIMP-1 mRNA expression was significantly reduced by transfecting them into the T6. CONCLUSION: We successfully constructed two TIMP-1 siRNA eukaryotic expression vectors and the transfected cells can significantly suppress the TIMP-1 expression.