登革病毒在白纹伊蚊细胞内免疫的研究

Research on intracellular immunization to dengue virus in the mosquito,Aedes albopictus cells

目的 研究白纹伊蚊对登革病毒产生细胞内免疫现象的分子机制。方法 扩增与克隆登革病毒NGC株的前膜蛋白基因prM ,将prM基因以 3种不同表达形式重组入昆虫表达载体pBh spEGFP ,以 3种重组质粒分别转染白纹伊蚊C6 36细胞 ,测定转染后的细胞对登革病毒感染的免疫效果。结果 构建了包含prM基因的 3种昆虫表达载体 ,Westernblot和荧光显微镜观察证明在C6 36细胞中成功表达了prM EGFP融合蛋白。MTT实验和空斑形成实验反映了C6 36细胞对登革病毒产生了细胞内免疫。结论 3种重组质粒均可诱导细胞内免疫现象 ,说明无论是否表达prM蛋白 ,只要有prM基因转录就可引起细胞内免疫现象 ;prM基因正义和反义转录形式均可诱导细胞内免疫 ;其形成机制可能是通过RNA干扰作用而产生。

Objective To research on the molecular mechanism of intracellular immunization to dengue virus in the mosquito cells. Methods prM gene of dengue virus was amplified and inserted into i nsect expression vector pB[hspEGFP] by three kinds of expression ways. The rec ombinant plasmids were used to transfect C6/36 cell. The phenomena of intracellu lar immunization of C6/36 cell against dengue virus were identified. Results Insect expression vectors including prM gene were constructe d. The expression of prM-EGFP fusion protein in C6/36 cell was tested by Wester n blot and fluorescent analysis. MTT and plaque formation test on C6/36 cell sho wed intracellular immunity to dengue virus. Conclusion Th e result showed that the transcription of prM gene caused intracellular immuniza tion, whether the prM protein was expressed or not. And intracellular immunizati on was induced by both sense and anti-sense transcription of prM gene. We infer red that the mechanism of intracellular immunization might be caused by RNA inte rference.