人乳头瘤病毒18型L1蛋白在昆虫杆状病毒系统中的表达  

Expression of L1 Protein of Human Papillomavirus Yype 18 in Baculovirus Expression System

作　　者：麻粉莲[1] 张骞[1] 郑丽舒[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所,北京100052

出　　处：《生物技术通讯》2015年第6期763-766,共4页Letters in Biotechnology

摘　　要：目的:利用昆虫杆状病毒系统表达人乳头瘤病毒18型(HPV18)L1蛋白。方法:将L1基因与p Fast Bac1(p FB1)载体连接,构建转移载体p FB1-L1,转化含Bacmid的大肠杆菌DH10Bac感受态细胞,获得穿梭质粒r Bacmid-L1;r Bacmid-L1转染Sf9细胞,获得重组病毒r Bac-L1;PCR法检测重组杆状病毒基因组L1基因,间接免疫荧光法和Western印迹检测L1蛋白的表达。结果:穿梭质粒r Bacmid-L1经PCR鉴定构建正确;感染r Bac-L1的Sf9细胞经PCR扩增可见1629 bp的特异性条带,间接免疫荧光检测可见绿色荧光,Western印迹鉴定与小鼠抗L1单克隆抗体发生特异性反应,在相对分子质量约60 000处可见特异性条带。结论:利用昆虫杆状病毒表达系统表达了HPV18 L1蛋白。Objective：To express the L1 protein of human papillomavirus type 18（HPV18） in baculovirus expression system.Methods：L1 gene of HPV18 was inserted into vector pFastBac1（pFB1）.The constructed transfer vector pFB1-L1 was transformed to competent E.coli DHlOBac cells containing Bacmid,and the obtained shuttle plasmid rBacmid-L1 was transfected into Sf9 cells,based on which recombinant baculovirus rBacmid-L1 was obtained and determined for L1 gene in genome by PCR,and for expression of L1 protein by IFA and Western blot.Results：PCR proved that shuttle plasmid rBacmid-Ll was constructed correctly.The gene at a length of1629 bp was amplified by PCR from,and green fluorescence was observed in the Sf9 cells infected with rBacmidL1.Western blot showed specific reactions of expressed product with mouse monoclonal antibody against HPV18L1 protein,which showed a band with relative molecular mass of about 60 000 Da.Conclusion：L1 protein of HPV18 was successfully expressed in baculovirus expression system.

关 键 词：L1蛋白 人乳头瘤病毒18型 杆状病毒 SF9细胞 

分 类 号：Q78[生物学—分子生物学]