Tat/DNA/Liposome复合物促基因转染的体外实验  

Tat/DNA/Liposome compound enhanced gene transfection in vitro

作　　者：周志益[1] 任建丽[2] 刘茜[1] 

机构地区：[1]重庆市第三人民医院老年病科 [2]重庆医科大学附属第二医院超声科,重庆400010

出　　处：《激光杂志》2013年第4期130-131,共2页Laser Journal

基　　金：国家自然科学基金青年科学基金项目资助课题(基金编号81000621);教育部博士点(新教师类)基金项目资助课题(基金编号20105503120008)

摘　　要：目的:探讨HIV-1 Tat蛋白转导域/质粒DNA/Liposome(TDL)复合物提高增强型绿色荧光蛋白质粒(pEGFP)在体外培养肝癌细胞株HepG2中转染效率的可行性。方法2?l Lipofectamine 2000加入以不同电荷比混匀的Tat肽与质粒DNA溶液中制成TDL复合物。琼脂糖凝胶电泳法观察Tat肽与质粒DNA的结合力;荧光显微镜和流式细胞仪观察TDL复合物、HIV-1 Tat肽与Lipofectamine 2000介导质粒DNA体外转染HepG2的效果;MTT法分析质粒DNA转染对HepG2活性的影响。结果琼脂糖凝胶电泳显示,各TDL复合物组均未发现明显DNA条带。TDL复合物的转染率优于单用Lipofectamine 2000(P<0.05),HIV-1 Tat肽未发生基因转染。DNA/Tat电荷比为1 8时,TDL复合物的转染效率最高。与空白组比较,各TDL复合物组、HIV-1 Tat肽组对细胞活性无明显影响(P>0.05),Lipofectamine 2000组降低细胞活性(P<0.05)。结论:TDL复合物可明显提高体外基因转染HepG2效率,该方法为提高非病毒载体的转染效率提供一个新策略,可能成为肝癌基因治疗的有效手段之一。Objective：To investigate the possibility of Tat/DNA/Liposome compound enhancing transfeetian efficiency of plasmid vector coding enhanced green fluorescence protein （pEGFP） in hmnan hepatoma cell line HepG2 ceils. Methods Tat peptide was mixed with Plasmid DNA by various charge ratio, then added 2.9 1 Lipofectamine 2000 to form TDL compound. Bonding force of Tat peptide and plasmid DNA was analyzed by agarose gel electrophoresis. Transfection effect of TDL compound,HIV - 1 Tat and Lipofectamine 2000 in HepG2 was observed by fluorescent microscopy and Fluorescence Activated Cell Scan. The viability of HepG2 was measured by MTr assay. Results A visible band was not found in TDL compound groups by agarose gel electrophoresis. Transfection efficiency of TDL compound was higher than that of Lipofectamine 2000 （P.9 0.05）. No transfection was happened in HIV- 1 Tat. When charge ratio of DNA and Tat was 1：8, transfection efficiency of TDL compound was the highest. There was no significant difference in cell viability between TDL groups and control group（P 〉 0.05）, while significant difference between Lipofectamine 2000 and control group（P？ 0.05）. Conclusion TDL compound can erahance gene transfection efficiency. This method offers a new strategy to increase the transfection efficiency of non - viral genetic vector, and it may be a useful tool for gene therapy of hepatocellular carcinoma （HCC）.

关 键 词：TDL复合物 肝癌细胞株HEPG2 基因转染 

分 类 号：TN248.1[电子电信—物理电子学]