以人乳头瘤病毒16 L1为载体的流感通用疫苗初步研究

目的 构建以人乳头瘤病毒(human papillomavirus,HPV)16 L1为载体的甲型流感病毒M2基因胞外区(M2e)通用疫苗杆粒,利用昆虫细胞杆状病毒表达系统,进行初步的蛋白表达.方法 利用PCR技术将甲型流感病毒M2e基因序列与HPVl6 L1相连.经酶切、酶联将融合基因插入pFastBacHTA载体,在DH10Bac细胞中进行同源重组,经测序鉴定后构建M2e-HPV16 L1杆粒,脂质体转染sf9昆虫细胞,收获并扩增含有M2e-HPV16 L1的杆状病毒,经扩增后获得高效价的重组杆状病毒,用SDS-PAGE、免疫荧光法和电镜检测目的蛋白的表达.结果 构建了以HPV16 L1为载体的M2e通用疫苗杆粒,通过转染sf9昆虫细胞得到初步M2e-HPV16 L1融合蛋白表达.结论 成功构建了HPV16 L1与甲型流感病毒M2e融合蛋白的病毒样颗粒,为流感通用疫苗的研制奠定了基础.

Research of universal influenza virus vaccine conjugated human papillomavirus 16 LI as a vector

Objective To construct a universal influenza virus vaccine Baemid containing M2 extracellular domain(M2e) of influenza virus which conjugated human papillomavirus(HPV)16 L1 as a vector and expressed the protein in insect baeulovirus system. Methods HPV16 L1 gene was amplified by PCR with the primers contained M2e gene. The conjugated genes were inserted into pFast HTA vector to recombine in DHI0Bac. The recombinant baemid was transfected into sf9 insect cells by liposome to produce baculovirus contained M2e-HPVI6 L1. Protein was determined by SDS-PAGE, immunofluorescence and electronmicroseope. Results The universal influenza virus vaccine Bacmid containing M2e of influenza virus which conjugated HPV16 L1 as a vector was successfully constructed. The recombinant protein wasexpressed in insect cells. Conclusions The recombinant protein was expressed in insect cells in the form of virus-like particles through the baculovirus system,which was a base of the universal influenza virus vaccine development.