摘要
目的获得有效表达人乳头瘤病毒16型(HPV16)L1基因的重组杆状病毒和腺病毒,为研究HPV的免疫保护机制提供材料。方法按照昆虫细胞密码子偏爱优化并合成HPV16L1基因,利用Bac-to-Bac昆虫表达系统获得表达HPV16L1基因的重组杆状病毒,利用AdEasy腺病毒载体系统获得表达HPV16L1基因的重组腺病毒载体。通过间接免疫荧光和Western blot对HPV16L1基因表达进行鉴定,利用负染电子显微镜观察病毒样颗粒(VLP)的形成。结果获得了稳定表达HPV16L1蛋白的重组杆状病毒和重组腺病毒载体,在Sf9细胞和293细胞中可有效表达能被抗HPV16L1单克隆抗体识别的L1蛋白,分子质量单位为56ku,在Sf9细胞中可观察到VLP的形成。结论按照昆虫细胞密码子偏爱进行优化的HPV16L1基因,在昆虫细胞和哺乳动物细胞内均可有效表达。
Objective To generate recombinant baculovirus and adenovirus which can express human papillomavirus type 16 （HPV16） L1 gene to provide materials for investigating the immunological mechanisms of HPV infection. Methods The HPV16L1 gene was artificially synthesized after optimization following insect cell codon bias. The recombinant baculovirus and adenovirus harboring the HPV16L1 were generated through Bac-to-Bac Baculovirus Expression System and AdEasy Adenoviral Vector System, respectively. The expression of the HPV16L1 was confirmed by both indirect immunofluorescence and Western blot. The virus-like particles （VLPs） of HPV16 was identified by negative staining electron microscopy. Results Both the recombinant baculovirus and adenovirus bearing HPV16L1 were successfully generated. The effective expression of HPV16L1 in insect Sf9 cells or 293 cells were detected by the monoclonal antibody which is specific to HPV16 L1, respectively, and the assembly of VLPs was observed in Sf9 cells. Conclusion The HPV16L1 gene optimized according to insect cell condon bias can be expressed effectively in both insect and mammalion cells.
出处
《中国病原生物学杂志》
CSCD
2007年第4期247-251,共5页
Journal of Pathogen Biology
关键词
人乳头瘤病毒u
16型
重组腺病毒
重组杆状病毒
表达
密码子优化
病毒样颗粒
Human papillomavirus type 16 L1
recombinant adenovirus
recombinant baculovirus
expression
codonoptimization
virus-like particle （VLP）
