Xie Na, Wang Xiao-yan, Zhang Qiong, Lin Yuan-yuan, Lin Ju-sheng
Institute of Liver Diseases, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Zhonghua Gan Zang Bing Za Zhi. 2006 Apr;14(4):268-71.
To construct four expression vectors carrying enhanced green fluorescent protein (EGFP) gene under the control of different HBV promoters, and to detect and analyze their expressions in hepatoma cell lines.
Four HBV promoters were amplified using PCR, and they were inserted into the T-vector and identified using restriction enzymes and sequencing, then cloned into the expression vector pEGFP-1. The four recombinant plasmids were transfected into human hepatoma cell line HepG2 by lipofectamine2000, and the positive cell clones were detected using fluorescence microscopy.
All target fragments were separately obtained and successfully cloned into the expression vector. The expressions of EGFP under the control of the four promoters were detected. The expressions of EGFP controlled by different promoters had some differences.
Reporter gene EGFP under the control of four HBV promoters can be specifically expressed in hepatoma cell line HepG2, and different promoters give different results; this may provide another option in gene therapy of liver diseases.
构建4种在不同乙肝病毒（HBV）启动子控制下携带增强型绿色荧光蛋白（EGFP）基因的表达载体，并检测和分析其在肝癌细胞系中的表达情况。
采用聚合酶链反应（PCR）扩增4种HBV启动子，将其插入T载体，用限制性内切酶和测序进行鉴定，然后克隆到表达载体pEGFP-1中。用脂质体2000将4种重组质粒转染人肝癌细胞系HepG2，用荧光显微镜检测阳性细胞克隆。
分别获得所有目的片段并成功克隆到表达载体中。检测了4种启动子控制下EGFP的表达情况。不同启动子控制下EGFP的表达存在一定差异。
4种HBV启动子控制下的报告基因EGFP可在肝癌细胞系HepG2中特异性表达，不同启动子产生不同结果；这可能为肝病基因治疗提供另一种选择。
