· 基础论着· ns2tp基因启动子报告基因载体的构建及其活性验证.pdf

· 332 · 中华实验和临床感染病杂志(电子版) 2013年6月第7卷第3期Chin J Exp Clin Infect Dis (Electronic Edition), June 2013, Vol. 7, No. 3 · 基础论著 · NS2TP基因启动子报告基因载体的构建及其活性验证 高学松 杨彪 王琦 李炜 成军 【摘要】 目的 研究丙型肝炎病毒非结构蛋白2反式调节基因 （NS2TP ）的转录调节机制。方 法 应用PCR方法扩增N S2TP 的启动子，克隆入pGL4.10载体，构建萤虫素酶报告基因载体，转染 HepG2细胞，检测双萤虫素酶活性，验证其转录活性；通过缺失突变法构建系列截短的N S2TP基因 启动子报告基因载体。结果 成功构建了系列截短的N S2TP基因启动子报告基因载体，分别命名为 N1 、N2 、N3和N4 ，并确定了N3 为NS2TP基因的核心启动子。结论 构建NS2TP基因启动子的系列截 短报告基因载体，确定了其最小转录活性区域，为进一步研究NS2TP基因的转录活性调节机制奠定理 论基础。 【关键词】NS2TP基因；报告基因载体；启动子 Construction of series reporter plasmids with truncated HCV NS2TP gene promoter GAO Xue-song*, YANG Biao, WANG Qi, LI Wei, CHENG Jun. *Department of General Medicine, Beijing Ditan Hospital, Capital Medical University, Beijing 100015, China Corresponding author: CHENG Jun, Email: chengjdt@ Abstract Objective To construct series of reporter plasmids with truncated NS2TP gene 【 】 promoter. Methods Fragments of NS2TP gene promoter was amplified by PCR and cloned into pGL4.10- Basic. Then the luciferase reporter vectors of NS2TP gene was constructed. Dual luciferase assays were performed with lysates of HepG2 cells transfected with NS2TP gene promoter reporter plasmids. Results Series of luciferase reporter plasmids with truncated NS2TP gene promoter were successfully constructed and named N1, N2, N3 and N4, respectively. N3 as the minimal promoter was determined. Conclusions Series of luciferase reporter plasmids with truncated NS2TP gene promoter were successfully constructed and their promoter activity were verified. These plasmids provide necessary experimental materials for further investigation of regulation of NS2TP gene. Key words 【 】 NS2TP gene; Reporter plasmid; Promoter activity 1丙型肝炎病毒 （hepatitis C virus，HCV ）属黄 端到羧基端依次为：结构蛋白Core 、E 1、E2和非 病毒科，大多数人感染HCV后表现为持续性感染，