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目的构建含有我国登革2型/4型病毒株嵌合E基因片段的真核表达质粒,观察重组质粒DNA的免疫原性,为登革多价疫苗的研究提供依据。方法首先将包含我国登革2型病毒43株E蛋白I/II抗原区和4型病毒B5株E蛋白III抗原区的嵌合E基因片段克隆至真核表达载体pcDNA3.1中,通过测序确定嵌合基因序列的正确性。然后将重组质粒以肌肉注射途径,免疫Balb/C小鼠。通过间接免疫荧光法对采集的鼠血清中的病毒特异抗体进行检测。结果构建的含有我国登革2型/4型病毒株嵌合E基因的真核表达质粒pcDNA-D2/4,经序列测定表明,导入的嵌合E基因片段的序列是正确的。将重组质粒DNA免疫小鼠,在初次免疫后的第3周,可同时检测到针对登革2型和4型病毒的特异荧光。结论所构建的含有不同血清型的我国登革病毒株嵌合E基因片段的真核重组质粒,可诱导小鼠同时产生针对两个血清型病毒的特异抗体,该研究为新型登革多价疫苗的研制提供了依据。
Objective To construct a eukaryotic expression plasmid containing the chimeric E gene fragment of dengue type 2/4 strain in China and observe the immunogenicity of the recombinant plasmid DNA to provide basis for the study of dengue multivalent vaccine. Methods The chimeric E gene fragment containing 43 E protein I / II antigen regions of Dengue 2 virus and E protein III antigen region of B4 virus strain was cloned into eukaryotic expression vector pcDNA3.1 and sequenced. Correctness of chimeric gene sequences. Balb / C mice were then immunized with the recombinant plasmid by intramuscular injection. The virus-specific antibodies in the collected mouse serum were detected by indirect immunofluorescence. Results The constructed eukaryotic expression plasmid pcDNA-D2 / 4 containing chimeric E gene of dengue type 2/4 strain in China showed that the sequence of the inserted chimeric E gene fragment was correct. The recombinant plasmid DNA was immunized in mice, the first three weeks after the first immunization, can detect specific Dengue 2 and 4 virus-specific fluorescence. Conclusion The eukaryotic recombinant plasmids containing different serotypes of chimeric E gene fragments of Dengue virus strains in China can induce specific antibodies against two serotypes in mice simultaneously. This study is a new dengue multivalent vaccine Provided the basis for the development.