转座子介导的多角体基因表达及提高病毒粒子的感染效率
Expression and package of polyhedrin gene of Bombyx mori nucleopolyhedrovirus by transposon
陈 璐1；李 兵2；贡成良2；吴 岩1；王文兵
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作者单位：1江苏大学生命科学研究院 镇江 212013； 2苏州大学生命科学学院 苏州 215123
中文关键词：家蚕核型多角体病毒，多角体基因，piggyBac，转基因，家蚕细胞
英文关键词：BmNPV,polyhedrin gene,piggyBac,transgenic,BmN cells
中文摘要：现行的杆状病毒表达外源基因的方法是将外源基因取代病毒中的多角体基因，因而得到的重组杆状病毒感染活体时不能经口感染，只能进行针刺注射，效率低且易引起活体感染其他疾病。将家蚕核型多角体病毒（Bombyx mori nucleopolyhedrovirus，BmNPV）中的多角体基因（polyhedrin,poly）及其启动子片段克隆到转座子载体pigA3GFP中，将其与辅助质粒pHA3PIG利用脂质体介导法导入家蚕细胞中，经过多次筛选获得稳定的转基因家蚕细胞。之后先将BmPAK6（含Lac Z）及BmGFP （含GFP）重组病毒分别感染转基因细胞，再将得到的重组病毒经口感染5龄家蚕幼虫。结果显示，重组杆状病毒可以经口感染家蚕幼虫。这些研究表明来自于转基因家蚕细胞的poly基因表达产物可以提高重组杆状病毒经口感染家蚕率，为解决杆状病毒表达系统中重组病毒不能经口感染家蚕幼虫的问题提供新思路。
英文摘要：The principle of baculovirus expression system is that substitute exogenous gene for polyhedrin gene (poly),and the recombinant baculoviruses lack the ability to infect insect larvae by oral inoculation.In order to deal with this problem,we cloned the poly gene with its promoter of Bombyx mori nucleopolyhedrovirus (BmNPV) into pigA3GFP vector,and transported it into BmN cells by lipofectamine with helper vector pHA3PIG.After several rounds of selecting,the transgenic BmN cells were obtained.Then the poly gene negative viruses,BmPAK6 (Lac Z) and BmGFP (GFP),infected the transgenic BmN cells.Subsequently,the viruses (BmPAK6 and BmGFP) from infected cells were used to orally inoculated the 5th instar larvae of Bombyx mori (B.mori),respectively.The results showed that B.mori larvae could be infected per os with the recombinant baculoviruses BmPAK6 and BmGFP,respectively.These results suggest that the products of poly gene expressed in the transgenic BmN cells could raise the pathogenicity of the recombinant virus in orally infected B.mori arvae.