密码子优化的鸭坦布苏病毒E基因在昆虫细胞中的表达  

EXPRESSION OF DUCK TEMBUSU VIRUS E GENE IN INSECT CELLS WITH CONDON OPTIMIZATION

作　　者：肖亚莉[1] 刘建龙[1] 李雪松[1] 余磊[1] 滕巧泱[1] 闫丽萍[1] 李国新[1] 李泽君[1] 

机构地区：[1]中国农业科学院上海兽医研究所,上海200241

出　　处：《中国动物传染病学报》2013年第3期13-17,共5页Chinese Journal of Animal Infectious Diseases

基　　金：国家自然科学基金项目(31172332);上海市科委基础研究重点项目(12JC1410600);上海市科技兴农重点攻关项目(沪农科攻字[2012]第2-6号);中央级公益性科研院所基本科研业务费(2011JB02)

摘　　要：为了使鸭坦布苏病毒(Duck tembusu virus,DTMUV)的E蛋白在Bac-to-Bac杆状病毒/昆虫细胞表达系统中高效表达,本研究将DTMUV的E基因按照昆虫细胞偏嗜性密码子进行了优化,合成了基因optiE,并将其插入到pFastBac HT载体上,得到质粒pFastBac-optiE。将该质粒转化DH10BAC感受态细胞,经抗性和蓝白斑筛选,获得重组转座子rBacmid-optiE。将rBacmid-optiE转染对数生长期的Sf9昆虫细胞,间接免疫荧光试验和Western blot方法都证实目的蛋白在重组杆状病毒感染的Sf9昆虫细胞中获得了表达。本研究为进一步研究DTMUV E蛋白的结构与功能奠定了基础。To express E protein of Duck tembusu virus（DTMUV）,E gene was optimized according to the bias codon usages of insect cells.The optimized E gene（opti-E） was cloned into the multiple cloning site of pFastBacHT vector of the Bac-to-Bac expression system.The recombinant plasmid pFastBac-optiE was then transformed into DH10Bac for blue/white selection and antibiotics selection.The positive recombinant bacmid rBacmid-optiE was transfected to Sf9 in logarithmic growth phase for harvesting recombinant baculovirus.The E protein expressed in Sf9 was confirmed in Western blot and IFA.The expression of E protein optimized was based for the further study of its structure and its function.

关 键 词：鸭坦布苏病毒 E蛋白 杆状病毒-昆虫细胞表达系统 密码子优化 

分 类 号：S852.659.6[农业科学—基础兽医学]