甲3型流感病毒M2基因在痘苗病毒天坛株中的表达

为获得表达甲3型流感病毒(H3N2)M2蛋白的重组天坛株痘苗病毒RVJ1175M2,使用PCR方法扩增流感病毒全长M2基因,将其克隆到天坛株痘苗病毒同源重组质粒pJSC1175中,获得重组质粒pJSC1175M2,通过与痘苗病毒载体同源重组,构建了含流感病毒M2基因的重组痘苗病毒株RVJ1175M2。PCR检测结果证明,流感病毒(H3N2)M2蛋白基因准确插入到天坛株痘苗病毒TK区;Western blot、免疫荧光和流式细胞计数表明重组病毒RVJ1175M2可以有效地表达M2蛋白,表达的M2蛋白有两条带,分别为15kD和13kD,与相关文献报道一致;M2蛋白可有效分布在感染细胞的细胞膜上。这些结果表明重组痘苗病毒株RVJ1175M2可以有效地表达流感病毒M2蛋白,为使用表达M2蛋白的不同类型疫苗进行广谱流感疫苗效果的比较研究奠定了基础。

Expression of Influenza A3 Virus (H3N2) M2 Gene in Vaccinia Virus Tiantan Strain

To construct a recombinant vaccinia virus RVJ1175M2 expressing influenza A3 virus(H3N2) M2 gene,full length gene encoding influenza virus(H3N2) M2 protein was amplified with PCR and cloned into plasmid pJSC1175 which was used for homologous recombination with vaccinia virus Tiantan strain.Along with this,a recombinant vaccinia virus RVJ1175M2 containing the M2 gene was subsequenlty constructed.It was identified by PCR that the gene of M2 protein was inserted into the TK locus of vaccinia virus Tiantan strain correctly and M2 protein was expressed by recombinant vaccinia virus RVJ1175M2 effectively.Two electrophoretic bands of M2 protein expressed by the infected HeLa cells,one of 15kD and the other of 13kD in accordance with related documents,was deteced by Western-blot.M2 protein distributing on the surface of the infected cells was demonstrated by immunofluorescence and flow cytometry.The results suggested that recombinant vaccinia virus RVJ1175M2 could express M2 protein effectively,this laid a foundation for comparative research on the immune effect of universal vaccine of influenza virus with different kinds of vaccine expressing M2 protein.