SARS病毒Spike蛋白重组腺病毒疫苗的构建及其免疫学分析  

Construction and immunogenicity of the recombinant adenovirus-SARS coronavirus(SARS CoV) spike glycoprotein

作　　者：郑尚永[1] 张冬梅[1] 曹毅[1] 张青峰[1] 沈璐辉[1] 潘卫庆[1] 

机构地区：[1]第二军医大学病原生物学教研室,上海200433

出　　处：《中国病原生物学杂志》2009年第5期329-332,336,F0002,共6页Journal of Pathogen Biology

摘　　要：目的构建基于SARS病毒Spike(S)蛋白的重组腺病毒,并对该重组腺病毒进行初步鉴定和免疫学探讨。方法通过全基因合成得到全长S蛋白基因,利用Ad5腺病毒系统构建S蛋白基因的重组腺病毒,采用间接免疫荧光(IFA)试验和蛋白免疫印迹法(Western blot)对S蛋白的表达进行鉴定,再用重组腺病毒免疫小鼠,观察小鼠抗体诱生情况,从而对该重组腺病毒的免疫效果进行初步评价。结果PCR扩增重组腺病毒质粒外源片段,大小为800 bp,与预期值一致;Western blot和IFA结果表明,重组腺病毒(rAd-S)表达S蛋白能被马抗SARS血清识别;ELISA检测结果显示,rAd-S能刺激小鼠机体产生抗体。结论重组腺病毒系统成功表达了SARS病毒S蛋白,免疫小鼠产生S蛋白特异的抗体。Objective To construct the recombinant adenovirus-SARS coronavirus spike glycoprotein and to investigate its immune response in mice, adenovirus was employed as vector to express the S protein of SARS in 293 cells. Methods A synthetic spike gene was synthesized by asymmetric PCR. An Ad vector expressing the S gene was constructed, and the expression of recombinant protein was confirmed by Western blotting and IFA analysis. To evaluate the immune responses of the recombinant adenovirus, BALB/c mice were immunized with the recombinant adenovirus at two doses. The serum antibody titers were measured by ELISA. Results PCR analysis showed that the 800 bp fragment amplified from the recombinant adenovirus plasmid was identical to the expected length. Western blot and IFA analysis indicated a detectable expression of the S protein of SARS-CoV （rAd-S）, and the S protein could be recognized by horse anti-serum against SARS cronavirus. Immunogenicity analysis also showed that rAd-S could elicit high titers of antibodies against SARS eoronavirus in mice. Conclusion The S protein was successfully expressed in adenovirus, and the expressed protein could induce antibodies specific for the S protein.

关 键 词：SARS SPIKE蛋白 重组腺病毒 疫苗 

分 类 号：R373.1[医药卫生—病原生物学]