Abstract: Objective　To perform prokaryotic expression and preliminary characterization of the recombinant poly⁃epitope vaccine EgG1Y162⁃2 (4) against cystic echinococcosis. Methods　The recombinant poly⁃epitope vaccine EgG1Y162⁃2 (4) against Echinococcus granulosus based on the linker GSGGSG was subjected to structural three⁃dimensional (3D) modeling using immunoinformatics to analyze the structural changes and evaluate the antigenicity of the vaccine. The pET30a⁃EgG1Y162⁃2 (4) recombinant plasmid was generated using double digestion with EcoR I and Sal I, and then transformed into competent cells. Following protein induction with isopropyl⁃β⁃D⁃thiogalactoside (IPTG), the prokaryotic expression proteins were characterized using Western blotting, and the antigenicity of the recombinant protein was analyzed using sera from cystic echinococcosis patients and health volunteers. Results　The four EgG1Y162⁃2 proteins coupled by the 3D structure of the recombinant vaccine EgG1Y162⁃2 (4) presented independent and effective expression and good antigenicity. The highest protein expression was detected in the supernatant following induction of the recombinant plasmid pET30a⁃EgG1Y162⁃2 (4) by 0.2 mmol/L IPTG at 37 ℃ for 4 h, and a pure protein component was seen following elution with 60 mmol/L imidazole. Western blotting analysis of the recombinant multi⁃epitope protein HIS⁃EgG1Y162⁃2 (4) showed a band at approximately 39 kDa, and this band was recognized by sera from cystic echinococcosis patients. Conclusion　A recombinant poly⁃epitope vaccine EgG1Y162⁃2 (4) against cystic echinococcosis has been successfully constructed, which provides a preliminary basis for researches on recombinant multi⁃epitope vaccine against cystic echinococcosis.

Key words: Cystic echinococcosis, Linker sequence, Multi?epitope vaccine, EgG1Y162?2 (4), Prokaryotic expression, Characterization

摘要： 目的　对细粒棘球蚴病重组多表位疫苗EgG1Y162⁃2（4）进行原核表达并初步鉴定。方法　利用免疫信息学方法对基于接头序列“GSGGSG”的细粒棘球蚴病重组多表位疫苗EgG1Y162⁃2（4）进行三维结构建模，分析其结构变化并评估该疫苗抗原性。通过EcoR I和Sal Ⅰ双酶切构建pET30a⁃EgG1Y162⁃2（4）重组质粒，将重组质粒转化入感受态细胞，利用异丙基⁃β⁃D⁃硫代半乳糖苷（isopropyl⁃β⁃D⁃thiogalactoside, IPTG）诱导蛋白表达， Western blotting鉴定原核表达蛋白产物，并应用细粒棘球蚴病患者及健康人血清分析重组蛋白抗原性。 结果　构建的重组疫苗EgG1Y162⁃2（4）三维结构串联的4个EgG1Y162⁃2蛋白均能独立、有效表达且具有良好抗原性。重组质粒pET30a⁃EgG1Y162⁃2（4）在0.2 mmol/L IPTG 37 ℃诱导4 h条件下，上清中目的蛋白表达量最高，使用60 mmol/L咪唑洗脱的蛋白成分较纯。Western blotting分析发现，重组多表位蛋白HIS⁃EgG1Y162⁃2（4）在约39 kDa位置处出现条带，且该蛋白可被细粒棘球蚴病患者血清识别。结论　成功构建了细粒棘球蚴重组疫苗EgG1Y162⁃2（4），为研究细粒棘球蚴病重组多表位疫苗奠定了基础。

关键词: 细粒棘球蚴病, 接头序列, 多表位疫苗, EgG1Y162?2（4）, 原核表达, 鉴定