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目的通过观察含我国登革2型病毒株D2-43的PrM-E基因的复制型SFV重组质粒DNA的免疫原性,为登革新型疫苗的研制提供依据。方法将PrM-E基因自T载体上切下,插入复制型SFV病毒载体质粒DNA中。将此重组质粒DNA以电穿孔法导入BHK21细胞,表达产物的特异性用间接免疫荧光法进行鉴定。采用去除内毒素的质粒提取试剂盒制备重组质粒DNA,然后以不同剂量通过肌肉注射途径免疫Balb/c鼠,鼠血清中的抗体用间接免疫荧光法进行检测。结果含PrM-E基因的重组SFV质粒DNA在BHK21细胞中可表达登革2型病毒的特异蛋白;经免疫Balb/c鼠后,鼠血清可与登革D2-43感染的C6/36抗原片起特异的抗原抗体反应。结论含登革2型病毒PrM-E基因的复制型SFV病毒载体质粒DNA在Balb/c鼠中可诱导登革2型病毒特异抗体的产生,但抗体水平较低。
Objective To observe the immunogenicity of recombinant plasmid DNA of SFV containing PrM-E gene of dengue type 2 strain D2-43 in China, and provide basis for the development of new dengue vaccine. Methods The PrM-E gene was excised from the T vector and inserted into the plasmid DNA of the replicative SFV vector. The recombinant plasmid DNA was electroporated into BHK21 cells, and the specificity of the expressed product was identified by indirect immunofluorescence. Recombinant plasmid DNA was prepared by removing the endotoxin plasmid extraction kit, and then Balb / c mice were immunized by intramuscular injection at different doses. The antibodies in the serum were detected by indirect immunofluorescence. Results Recombinant SFV plasmid DNA containing PrM-E gene could express dengue virus type 2 specific protein in BHK21 cells. After immunization of Balb / c mice, the serum of mice could be infected with C6 / 36 antigen From a specific antigen-antibody reaction. Conclusion Plasmid DNA of replicative SFV virus vector containing PrM-E gene of dengue 2 can induce dengue virus type 2 specific antibody production in Balb / c mice with low antibody level.