表达流行性感冒病毒血凝素蛋白的重组腺病毒的构建

Construction of Recombinant Adenovirus Expressing Influenza Virus Haemagglutinin

用RT PCR方法扩增流感病毒血凝素基因 ,将其克隆到 5型腺病毒载体质粒 pAd5C中。用此质粒与EcoRI酶切回收的 5型腺病毒基因组大片段共转染 2 93细胞 ,通过PCR初步筛选得到重组病毒。SDS PAGE电泳、West ernblot证明重组病毒能表达流感病毒的血凝素蛋白。此重组病毒经滴鼻免疫Balb/C小鼠 ,ELISA实验证明能诱导小鼠产生针对流感病毒的循环抗体IgG和分泌型抗体IgA。本实验证明 ,利用E3区缺失的 5型腺病毒载体可以用来表达流感病毒血凝素抗原 ,重组抗原具有良好的免疫原性 。

The HA gene of influenza virus A1/PR8/34 was amplified by RT-PCR and cloned into the shuttle plasmid pAd5C. The constructed plasmid and the largest fragment of adenovirus type 5 genome DNA digested by EcoR I were cotransfected into 293 cells. Recombinant virus was selected by PCR. The expression of HA gene in the cells infected with recombinant virus was identified by SDS-PAGE and Westen blot. Mice immunized with the recombinant virus by nasal route can produce HA specific antibody both in serum and on mucosal surface. The Ad5 vector with E3 region deletion can be used as a vector to construct the recombinant influenza virus vaccine.