【摘要】 构建以TK为基本启动子,其上游由抗氧化反应元件(ARE)调控的报告载体pARE-TK-GFP以期用于化学预防剂的筛选.应用重组PCR技术,分别经3次PCR获得重组TK启动子并克隆入pMD18-T载体中,经酶切和测序鉴定正确后连入pEGFP中构建载体pTK-GFP.人工合成的ARE插入TK基本启动子上游,成功构建报告载体pARE-TK-GFP.脂质体转染HepG2细胞并在荧光显微镜下观察有绿色荧光.该载体的成功构建可为各种天然或人工合成的化学预防剂的筛选奠定基础.更多还原

【Abstract】 A reporter vector under the transcriptional control of thymidine kinase(TK) promoter and the regulation of anti-oxidative response element(ARE) was constructed so as to screen the chemopreventive agents.The recombinant TK promoter was amplified by three times PCR and subcloned into pMD18-T vector then inserted into pEGFP after digested by restriction enzyme and sequencing to constructed pTK-GFP.The artificial synthetic ARE was inserted into pTK-GFP to construct the reporter vector pARE-TK-GFP.Finally the pARE-TK-GFP was transfected into HepG2 cells and the green fluorescence was observed under the fluorescent microscope.The results showed that this reporter vector was successfully constructed,which provides a basis for the screening of natural or synthetic cancer chemopreventive agents.更多还原