中文摘要：

为构建流行性乙型脑炎病毒（JEV） E蛋白多表位疫苗，本研究将人工合成的优化后E蛋白多表位肽（MP）序列克隆于酵母表达载体pPICZα-A中，构建分泌型重组酵母表达质粒pPICZ-MP后电转化导于毕赤酵母X-33中，重组酵母菌组子经甲醇诱导表达重组蛋白（rMP）。将其免疫小鼠后，通过测定rMP免疫小鼠的抗体水平、抗体亚型、中和抗体和CTL，以评价该多表位疫苗在小鼠模型的免疫原性。结果显示正确构建了表达JEV E蛋白多表位抗原的重组酵母菌株X33 （pPICZ -MP），并分泌表达多表位肽rMP ；纯化后的rMP免疫小鼠产生了较强的体液免疫和细胞免疫反应，其中和抗体水平、CTL活性均与活病毒疫苗组无显著差异（p〉0.05），而且能保护小鼠免受致死量JEV的攻击。上述结果表明本研究以酵母菌表达的JEV E蛋白多表位疫苗具有良好的的免疫原性，能够刺激小鼠产生特异性的细胞免疫和体液免疫应答反应，为流行性乙型脑炎多表位疫苗研制提供了新的思路。

英文摘要：

This study aimed to construct a multi-epitope vaccine of Japanese encephalitis virus （JEV） envelope protein and evaluate its immunogenicity and protective efficacy in mice model. In this study, A codon optimized multiple-epitope peptide gene （MP） from E protein of JEV was cloned into the plasmid pPICZα-A to construct the recombinant expression plasmid pPICZ-MP, which was linearized with SacⅠand transformed into Pichia pastoris X-33 by electroporation for expression. Then the recombinant multi-epitope peptide （rMP） were purified with Ni-NTP column from the recombinant P.pastoris X-33 culture medium. Mice were immunized with the rMP, and the immuno-responses were detected to evaluate the immunogenicity and protective efficacy of rMP in mice model. Animal experiments shown that the mice immunized with rMP had high humoral and cellular immune responses,and were protected against lethal JEV challenge. The neutralizing antibody level and CTL of mice immunized with the protein showed no significant difference with live JEV vaccine. The results indicated that the rMP was able to induce strong immune responses, suggesting that the rMP might be an promising subunite candidate vaccine for preventing JEV infection.