摘要
由于ＨＰＶ１６Ｅ６蛋白能诱导机体保护性免疫反应，可作为基因治疗的靶抗原。用杆状病毒昆虫细胞表达系统制备了ＨＰＶ１６Ｅ６基因工程蛋白，拟用于宫颈癌细胞系小鼠模型抗癌的免疫治疗。用ＰＣＲ技术从ＨＰＶ１６基因组中扩增获得转化基因Ｅ６的完整ＯＲＦ，按ＴＡ策略将其克隆到自行制备的杆状病毒转移载体ｐＶＬ１３９３Ｔ尾载体中，置于杆状病毒ＡｃＭＮＰＶＰｏｌｈ晚期启动子控制之下，用此重组转移质粒ｐＶＬ１３９３Ｅ６与杆状病毒ＤＮＡ共转染昆虫细胞Ｓｆ９，经噬斑筛选获得带有编码Ｅ６蛋白基因的重组杆状病毒株，并在昆虫细胞Ｓｆ９中表达为非融合性Ｅ６蛋白。ＳＤＳＰＡＧＥ电泳分析其分子量约为１８ｋＤ，免疫印迹实验表明，此重组蛋白能被兔抗ＨＰＶ１６Ｅ６抗体所识别。
The baculovirus insect cell expression system was used to prepare the HPV16 E 6 recombinant protein. The HPV16 E 6 complete ORF amplified by PCR from HPV16 genome was T A cloned into the baculovirus transfer vector pVL1393 T under AcMNPV Polh. This recombinant transfer plasmid pVL1393 E 6 DNA and baculovirus DNA were co transfected into insect cell Sf 9. After the plaque screening, the recombinant baculovirus AcMNPV E 6 was expressed in insect cells. The E 6, analysised by SDS PAGE, was 18kD, and was recognized by antibody against E 6 protein, which showed their natural antigenecity.
出处
《中国病毒学》
CSCD
1998年第4期300-305,共6页
Virologica Sinica
基金
山东省卫生厅科研基金
关键词
HPV16型E6
昆虫细胞
表达
人乳头瘤病毒
Human Papillomavirus Type 16 E 6, T A cloning, Baculovirus, Insect cell,Expression
