携带绿色荧光报告基因的EBV-LMP1真核表达载体构建与鼻咽癌细胞中的表达  

The Construction of the Eukaryotic Expression Vectors with the EBV-LMP1 Carrying Green Floprescent Report Genes and Expressed in Nasopharyngeal Carcinoma Cells

作　　者：撖子建 何芹[2] 鄢雪敏 闫琳琳[1] 蔡琰[2] 黄元姣[1,2] 

机构地区：[1]广西医科大学医学科学实验中心,南宁530021 [2]广西医科大学生物化学与分子生物学教研室,南宁530021

出　　处：《基因组学与应用生物学》2015年第6期1137-1143,共7页Genomics and Applied Biology

基　　金：国家自然科学基金(No.81260405);广西自然科学基金(No.2011GXNSFA018233);广西区域性高发肿瘤早期防治研究重点实验室基金项目(GK2013-A-02-02)共同资助

摘　　要：潜伏膜蛋白1(LMP1)是由EB病毒编码的致瘤蛋白,众多研究表明LMP1蛋白可通过NF-κB、p38 MAPK、c-JNK等多条重要信号通路引起鼻咽癌细胞的生物学行为改变。我们从EB病毒阳性的B95-8狨猴淋巴瘤细胞中克隆EB病毒LMP1 c DNA,构建携带绿色荧光基因的真核表达质粒p IRES2-Zs-Green1-LMP1,通过脂质体转染的方法将质粒导入鼻咽癌细胞株CNE1、CNE2中,利用质粒所携带的绿色荧光蛋白表达粗略计算转染的效率,通过免疫细胞化学(ICC)、RT-PCR、Western-Blot检测该质粒的表达。本实验室所构建的p IRES2-Zs-Green1-LMP1表达质粒能在鼻咽癌细胞内表达LMP1蛋白,为后续的实验研究奠定基础。The latent membrane protein 1(LMP1) is a tumorigenic protein encoded by EB viruses, numerous studies have shown that LMP1 protein can change nasopharyngeal carcinoma cells' biobehavioral through NF-kappa B,p38 MAPK, c-JNK and other important signaling pathway. We cloned the LMP1 c DNA from the EBV positive B95-8 cells to construct eukaryotic expression plasmid p IRES2-Zs-Green1-LMP1 plasmid which carrying green fluorescent gene, by transfecting plasmid into nasopharyngeal carcinoma cell lines CNE1, CNE2, the rough transfection efficiency was calculated by expression of green fluorescent protein, use Immune Cells Chemical(ICC), RT-PCR, Western-Blot to detect the LMP1 expressed. Constructed p IRES2 Zs-Green1-LMP1 expression plasmid by our laboratory could express LMP1 protein in nasopharyngeal carcinoma cells, and lay the foundation for subsequent experimental study.

关 键 词：EBV潜伏膜蛋白1(EBV-LMP1) c DNA克隆 绿色荧光蛋白真核表达载体 质粒p IRES2-Zs-Green1 

分 类 号：R739.63[医药卫生—肿瘤]