BmNPV多角体蛋白片段引导外源蛋白固定入多角体的研究  

Immobilization of Foreign Protein into BmNPV Polyhedra Through Fusion with Partial Polyhedrin Fragment

作　　者：曹翠平[1] 于少芳[1] 郭爱芹[1] 杨锐[1] 相兴伟[1] 陈琳[1] 吴小锋[1] 

机构地区：[1]浙江大学动物科学学院,杭州310029

出　　处：《蚕业科学》2010年第2期282-289,共8页ACTA SERICOLOGICA SINICA

基　　金：国家自然科学基金项目(No.30871827);浙江省科技厅国际合作项目(No.2009C14G2010086)

摘　　要：家蚕核型多角体病毒(BmNPV)晚期产生大量多角体,其主要作用是包埋病毒粒子免受外部恶劣环境的影响,为次代感染提供保障。将BmNPV多角体基因分段克隆并与报告基因——增强型绿色荧光蛋白(EGFP)基因融合,利用BmNPVPolh+Bac-to-Bac表达系统构建重组病毒,使多角体蛋白与融合蛋白在BmN细胞内共表达。分析感染细胞和形成的多角体中融合蛋白的含量,研究不同的多角体蛋白片段引导EGFP固定入多角体的效果,结果表明构建的5种重组病毒都能大量表达融合蛋白,每种多角体蛋白片段都能引导EGFP固定入多角体内部,且外源融合蛋白在多角体总蛋白中占有较高的比例,其中,以多角体蛋白N末端100个氨基酸和完整多角体蛋白序列作为信号固定EGFP的效率最高。这一现象为解决活性蛋白长期保存提供了新思路,同时对于开发含有毒素蛋白的新型生物杀虫剂和新型疫苗的转运载体也具有较好的参考价值。A large amount of polyhedra is produced in late phase of BmNPV infection to protect the incorporated virions from unfavorable environmental influences for the benefit of next generation infection. In this study, five different fragments of BmNPV polyhedrin gene were cloned and ligated with enhanced green fluorescent protein gene （egfp） as a reporter. Using BmNPV Polh ＋ Bac-to-Bac expression system, five recombinant baculoviruses were constructed to coexpress the fusion proteins with polyhedrin in BmN cell. The results showed that the fusion proteins were abundantly expressed in all 5 constructed recombinant baculoviruses and were able to guide the embedment of EGFP into polyhedra at a high ratio. Moreover, the foreign fusion protein took a fairly high percentage in the total polyedrin proteins. In particular, the fusion protein with 100 aa fragment at N-terminal and with full-length of polyhedrin presented best protein incorporation into polyhedra. This result provided a new inspiration for efficient preservation of active proteins and development of new biopesticide with toxin protein and new delivery vector system for vaccines.

关 键 词：家蚕核型多角体病毒 多角体 多角体蛋白片段 蛋白质固定 

分 类 号：S884.5[农业科学—特种经济动物饲养]