Construction and identification of matrix metalloproteinase inhibitor-1 siRNA eukaryotic expression vectors.
- Author:
Li TIAN
1
;
Ji-chang LI
;
Guo-qiang ZHAO
;
Kui-sheng CHEN
;
Wen-hui FAN
;
Xin LOU
;
Miao-miao SUN
;
Xue-quan CAO
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Cell Line; Gene Silencing; Genetic Vectors; Hepatic Stellate Cells; Plasmids; RNA, Small Interfering; Rats; Tissue Inhibitor of Metalloproteinase-1; genetics; Transfection
- From: Chinese Journal of Hepatology 2007;15(3):196-198
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct TIMP-1 siRNA eukaryotic expression vectors and evaluate their effect on TIMP-1 mRNA expression in hepatic stellate cells.
METHODSThe 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.
RESULTSTargeting sequences of TIMP-1 siRNA eukaryotic expression vectors were correct. TIMP-1 mRNA expression was significantly reduced by transfecting them into the T6.
CONCLUSIONWe successfully constructed two TIMP-1 siRNA eukaryotic expression vectors and the transfected cells can significantly suppress the TIMP-1 expression.
