猪流感复合多表位核酸疫苗的构建及其表达研究

目的构建猪流感病毒复合多表位核酸疫苗,并研究其表达产物的抗原性。方法以H3、H9亚型流感的HA抗原表位及流感病毒的其它主要抗原(NP、NA、M)表位基因为基础,再附以Kozak序列和适当的酶切位点,设计并合成一段长765bp的复合多表位基因(Epi),其中各表位基本独立,将其克隆入真核表达载体pIRES1neo,构建重组质粒pIRE-Epi。将多表位抗原基因Epi插入到融合表达基因H57中,构建重组质粒pIRE-H57-Epi。将构建的重组质粒在Hela细胞中表达,并用免疫荧光方法初步检测所表达蛋白的抗原性。结果所构建的融合表达载体pIRE-Epi和pIRE-H57-Epi,在Hela细胞中表达出流感病毒多表位抗原蛋白,并具有抗原性。结论已成功构建猪流感病毒多表位基因,有可能成为较理想的猪流感候选疫苗。

Construction and Expression of Porcine Influenza Virus Multi-epitope DNA Vaccine

Objective To construct porcine influen{a virus multi-epitope DNA vaccine and study the antigenicity of expressed product.Methods Design and synthesize a DNA fragment containing the HA antigenic epitopes of influenza virus subtype H3 and H9 as well as the epitopes of other major antigens(NP,NA and M) of influenza virus,Kozak sequence and proper restriction sites,in which the epitopes were separated by spacer sequence(-P-P-S-),and clone into eukaryotic expression vector pIRES1neo to construct recombinant plasmid pIRE-Epi.Clone multi-epitope antigen gene Epi into plasmid pUCM-T-H57 to construct a transfer vector pUCM-T-H57-Epi,then clone H57-Epi into pIRES1neo to construct expression vector pIRE-H57-Epi.Transformed the constructed recombinant plamids to Hela cells,and determine the antigenicity of expressed protein by IFA.Results Both fusion expression vectors pIRE-Epi and pIRE-H57-Epi were successfully constructed,and porcine influenza virus multi-epitope antigen with antigenicity was expressed in Hela cells.Conclusion Porcine influenza virus multi-epitope DNA was successfully constructed,which might be an ideal candidate for porcine influenza vaccine.