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目的 :构建我国登革 2型病毒 4 3株prM E基因的甲病毒 (Semlikiforestvirus ,SFV)重组RNA ,为登革新型疫苗的研究奠定基础。方法 :首先将含有多种稀有限制酶位点的接头插入pSFV载体的多克隆位点 ,再把prM E基因克隆至该载体中。将获得的重组质粒DNA经体外转录后 ,通过电穿孔法将其转录的RNA产物导入宿主细胞 ,并采用间接免疫荧光法检测prM E基因的表达。结果 :已获得含多种酶位点的pSFV病毒载体 ,并且所构建的含prM E基因的重组pSFV病毒RNA ,在宿主细胞中的表达产物可与登革 2型病毒特异抗体起反应。结论 :构建的含prM E基因的重组pSFV病毒RNA可表达我国登革 2型病毒株的特异蛋白。
OBJECTIVE: To construct the recombinant RNA of Semlikiforestvirus (SFV) of prM E gene of 43 strains of dengue 2 virus in our country and lay a foundation for the study of new dengue vaccine. Methods: First, a linker containing multiple restriction enzyme sites was inserted into the multiple cloning site of the pSFV vector and the prM E gene was cloned into the vector. The obtained recombinant plasmid DNA was transcribed in vitro and its transcribed RNA product was introduced into the host cell by electroporation, and the expression of prM E gene was detected by indirect immunofluorescence. Results: The pSFV viral vector containing multiple enzyme sites was obtained, and the constructed recombinant pSFV viral RNA containing the prM E gene was able to react with Dengue virus type 2 specific antibodies in host cells. CONCLUSION: The constructed recombinant pSFV RNA containing prM E gene can express specific proteins of Dengue 2 strain in China.