在昆虫细胞中表达西尼罗河病毒prM+E蛋白

Expression of West Nile virus prM + E protein in insect cells

采用PCR技术扩增prM+E基因,将其插入到供体质粒pFast-Bac1中得到重组质粒Pfast-Bac1-W-PE,之后转化大肠杆菌DH10-Bac,构建包含西尼罗河病毒prM+E基因的重组杆状病毒,然后感染正常的sf9细胞,收集感染不同时间段的细胞及上层培养基。结果表明:在感染后的第1天目的基因就得到了转录,同时有少量的重组杆状病毒释放到培养基中;SDS-PAGE和Western-Blot检测结果显示,在43.0ku与66.0ku条带之间,感染细胞裂解液较正常细胞的裂解液多出一条带,该蛋白为WNV完整的E蛋白,而且这一差异蛋白能与WNV兔多抗反应。

Thepr M＋Egene was amplified by PCR and inserted into the plasmid pFast-Bac1 to obtainrecombinant plasmid Pfast-Bac1-W-PE. The recombinant plasmid was transformed intoEscherichia coliDH10-Bac,and a recombinant baculovirus containing thepr M＋Egene ofWest Nile viruswas constructed. The normalSf9 cells were infected by the recombinant baculovirus,the cells and upper medium were collected at differenttime intervals. The results showed that thepr M＋Egene were transcribed in the first day after infection,and asmall amount of recombinant baculovirus was released into the medium. The results of SDS-PAGE and Western-Blot showed that between 43. 0—66. 0 ku,the lysate of infected cells was more than 1 strip of normal cell lysate.The protein was a complete E protein of WNV, and this differential protein could react with WNV rabbitpolyclonal antibody.