人乳头瘤病毒16L1/E7CTL重组杆状病毒载体的构建及鉴定

Construction and Identification of the Recombinant Bacmid of Human Papillomavirus 16 L1/E7CTL Gene

目的研制人乳头瘤病毒16型(HPV16)L1/E7CTL重组杆状病毒载体。方法以包含HPV16标准病毒株的质粒pHPV16为模板,采用PCR技术扩增HPV16主要衣壳蛋白L1(1-471aa)的基因序列,同时人工合成E7蛋白CTL表位E749-57的编码序列,将两个目的基因逐步连接构建重组质粒pUC19HPV16L1/E7CTL,HPV16L1/E7CTL片段经质粒pFast Bac1转位至Bacmid DNA后通过PCR鉴定重组杆粒。结果成功构建了包含融合基因HPV16L1/E7CTL片段的克隆质粒pUC19HPV16L1/E7CTL,制备了L1/E7CTL重组杆状病毒载体杆粒BACMID DNA。结论采用分子克隆技术构建了包含L1/E7CTL的重组杆粒BACMID DNA,有利于下一步融合蛋白的表达,制备一种能够更有效激发CTL杀伤作用的嵌合病毒样颗粒疫苗,为疫苗研制打下基础。

Objective To construct and identify a recombinant Baculovirus Bacmid DNA vector of Human Papillomavirus16（HPV16） L1/E7CTL. Methods Amplified major capsid protein L1 （1-471aa） gene sequence by Polyenzyme Chain Reaction （PCR）, taking standard virus pHPV 16 as a plate. The composed E749-57 coding gene was combined to L1 to construct recombinant plasmid pUC19HPV16L1/E7CTL HPV16L1/E7CTL sequence was transferred to Bacmid DNA by deliver plasmid pFast Bacl, and the recombinant Bacmid DNA was identified by PCR. Results Plasmid pUC19HPV16L1/ E7CTL including the combined genes and recombinant Bacmid DNA were constructed. Conclusion These data indicate that recombinant Bacmid DNA vector of HPV16L1/ETCTL was constructed. It is helpful to express recombinant protein and prepare a more effective chimeric virus-liked vaccin next step, and it also can be a foundation for HPV vaccines technology exploration.