应用水动力转染技术使人低密度脂蛋白受体、CD81和浸入诱导蛋白L同时在小鼠体内表达  

Co-Expression of Human LDLR CD81 and Sip-L Molecules in Mouse Using Hydrodynamics Transfection

作　　者：贾帅争[1] 胡锦辉[2] 郑晓玲[3] 吕丽萍[1] 刘敏霞 詹林盛[1] 王全立[1] 

机构地区：[1]军事医学科学院输血医学研究所,北京100850 [2]上海中医药大学附属曙光医院,上海200021 [3]军事医学科学院附属医院,北京100039

出　　处：《生物技术通讯》2005年第4期395-398,共4页Letters in Biotechnology

基　　金：国家自然科学基金项目(30300184);军事医学科学院创新基金项目

摘　　要：为建立人低密度脂蛋白受体(LDLR)、CD81和浸入诱导蛋白L(Sip-L)等多个HCV感染相关分子共表达的转基因小鼠,首先分别构建了白蛋白启动子调控的人LDLR、CD81和Sip-L基因的小鼠肝脏组织特异性表达载体,将3种质粒同时采用水动力转染技术导入小鼠体内,RT-PCR和免疫组化技术检测转入体内基因的表达及持续时间。结果表明,转染8h后即可检测到3种基因在体内转录,人LDLR和CD81在转染1～4d后可在50%～90%的肝细胞中高效表达,7d后检测不到目的基因表达。为了进一步延长转染基因在小鼠体内的表达时间,又分别构建了人LDLR、CD81和Sip-L的染色体整合型表达载体,将它们与噬菌体ФC31的整合酶表达载体同时水动力转染小鼠,3种基因在体内表达时间可持续到转染后25d。以上结果表明建立了多个HCV感染相关分子共表达的转基因小鼠,为确定这些分子能否使小鼠感染HCV奠定了基础。To establish a transgenic mouse which coexpressing human LDLR, CD81 and Sip-L, three eukaryotic expression vectors of human LDLR, CD81 and Sip-L gene under control of mouse liver tissue-specific albumin promoter were constructed, respectively. These vectors were transfected together into immunocompetent KM mice in vivo using hydrodynamics transfection method. Expression of the transfected genes were analyzed with immunohistochcmistry and RT-PCR method. Transcription of human CD81, LDLR and Sip-L genes were all detected at 8 h post transfection. Efficient expression of human LDLR and CD81 molecules were detected in about 50%-90% hepatocytes from 1-4 d post transfection. Expression of these three human genes lasted no more than 7 d. In order to make these genes express longer, other three eukaryotic expression vectors which all contained attachment site attB sequence of phage ФC31 integrase were constructed. Human LDLR, CD81 and Sip-L gene in these three vectors were all under control of mouse liver tissue-specific promoter. Then vectors of human LDLR, CD81 and Sip-L gene and the vector expressing phage ФC31 integrase were mixed together and transfected into mouse hepatocytes. Expression of human CD81 and LDLR lasted about 25 d in 50%-90% mouse hepatocytes. These results showed that transgenic mouse which coexpressing human CD81, LDLR and Sip-L were obtained. This is the basis to determine whether these three genes can confer mouse susceptibility to HCV infection.

关 键 词：丙型肝炎病毒 低密度脂蛋白受体 CD81 浸入诱导蛋白L 水动力转染 

分 类 号：Q786[生物学—分子生物学]