流感病毒血凝素基因HA1区的克隆及其真核表达载体的构建

Cloning of the HA1 region from gene encoding the influenza virus A hemagglutinin and construction of its eukaryotic expression plasmid

目的 克隆甲型流感病毒血凝素基因HA1区及构建其真核表达载体。方法 从接种人流感病毒株A/PR/ 8/ 34(H1N1)的鸡胚尿囊液中提取病毒RNA ,用特异引物进行RT PCR ,扩增血凝素基因HA1区。将所扩增片断克隆入真核表达质粒载体pcDNA3.1(+) ,转化感受态大肠杆菌JM 10 9并筛选阳性克隆。结果 经双酶切、PCR及测序鉴定证实血凝素基因HA1区的真核表达载体构建成功。结论 甲型流感病毒血凝素的HA1区是与宿主细胞膜表面受体结合的部位并起着诱导机体产生保护性抗体的作用 ,HA1区的克隆和其真核表达质粒的构建将为防治病毒侵入宿主细胞及核酸疫苗的研究打下基础。

To clone the HA1 region from gene encoding the influenza virus A hema agglutinin and to construct its eukaryotic expression plasmid, viral RNA was extracted from allantoic fluid of chicken embryos inoculated with influenza virus A /PR8/34, and the HA1 region gene was amplified by using specific primer and RT PCR.The amplified DNA fragment was then cloned into eukaryotic expression plasmid pcDNA3.1(+). The E.coli competent cells were transformed with recombinant plasmid and positive clones were selected.By restriction enzyme identification, PCR and sequence analysis, the recombinant plasmid of the HA1 was successfully constructed.It suggests that the HAI region of gene coding the hemagglutinin of influenza virus A is the major binding area with host cells and can induce the production of protective antibodies.