雌二醇依赖性的细胞表达载体的研究.pdfVIP

中国农业科学 2005,38(5):1046-1051 Scientia Agricultura Sinica 雌二醇依赖性的细胞表达载体的研究 杨桂香, 彭险峰, 曾振灵, 陈杖榴 （华南农业大学兽医学院/广东省兽药研制与安全评价重点实验室，广州 510642 ） 摘要：根据雌激素类化合物在机体内的作用机理，以增强型绿色荧光蛋白（EGFP）基因为报告基因构建了 4 个启动子不同的表达载体(pERE-SV-EGFP, pERE-TA-EGFP, pERE-CMV-EGFP, pERE-TK-EGFP)。以这些表达载体瞬时 转染 MCF-7 细胞，报告基因的基础表达与启动子的强度相关。以 17β-雌二醇(E -9 -1 2 ,10 mol·L )诱导表达载体瞬 时转染的 MCF-7 细胞，发现在表达载体 pERE-TA-EGFP、pERE-TK-EGFP、pERE-SV-EGFP 转染的 MCF-7 细胞中，E2 能诱导 2 倍以上的报告基因的表达。这表明在这些瞬时表达系统中报告基因的表达是雌激素依赖性的，因此这些 表达载体可用于稳定转染以建立稳定表达分析系统。 关键词： 雌激素类化合物; EGFP; 表达载体; 瞬时转染 Construction of Expression Vectors for Detection of β Estradiol YANG Gui-xiang, PENG Xian-feng, ZENG Zhen-ling, CHEN Zhang-liu (Guangdong Key Laboratory for Veterinary Pharmaceutics Development and Safety Evaluation/College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642) Abstract: The estrogenic endocrine disrupting compounds whether from environment and food chains or from chemicals illegally used in animal production pose a serious threat to the safety of animal derived products. Because of the huge diversities of these chemical structures, it is necessary to establish high-throughput screening and detecting methods to screen them. In this study, 4 recombination expression vectors with enhanced GFP as reporter gene were constructed. These constructs, namely, pERE-SV-EGFP, pERE-TA-EGFP, pERE-CMV-EGFP, pERE-TK-EGFP, only differ in promoters. Then the four expression vectors were transiently transfected into MCF-7 cells with liposome transf