传染性法氏囊病病毒VP5蛋白跨膜区的敲除及可溶性原核表达研究  

Knockout of IBDV VP5 Protein Transmembrane Region and the Soluble Prokaryotic Expression Research

作　　者：严孝金[1] 李锋[1] 秦立廷[2] 李倩倩[1] 韩翠晓[1] 冯舵[1] 王笑梅[2] 高伟[1] 

机构地区：[1]北京林业大学理学院,北京100083 [2]中国农业科学院哈尔滨兽医研究所,黑龙江哈尔滨150001

出　　处：《安徽农业科学》2011年第17期10461-10463,共3页Journal of Anhui Agricultural Sciences

基　　金：国家自然科学基金项目(30970578;31070651);教育部"新世纪优秀人才支持计划"项目(NECT-08-0731)

摘　　要：[目的]构建敲除传染性法氏囊病病毒(IBDV)VP5蛋白跨膜区序列的原核表达载体,并进行目的蛋白的表达、分离和纯化。[方法]利用PCR技术分别扩增IBDVVP5基因的胞外片段和胞内片段,然后将2个片段及pET-28b(+)同时相接,即载体-胞内片段-胞外片段-载体,构建了敲除跨膜区基因片段的VP5重组表达质粒pET-VP5-FC及改进后的pET-VP5-SC。将表达质粒转化BL21(DE3),IPTG诱导后经Ni亲和层析及凝胶过滤层析纯化重组蛋白。[结果]得到可溶性表达的IBDVVP5。[结论]为进一步研究VP5蛋白的结构与功能奠定了基础。[ Objective ] To construct the prokaryotic expression vector of IBDV VP5 protein whose transmembrane region sequence was knocked out. Moreover, the expression, sep^ation and purification of objective protein were carried out. [ Method] PCR technology was used to respectively amplify the extracellular and intracellular fragments of IBDV VP5 gene. Then, the two fragments were simultaneously linked to pET-28b （ ＋ ）, and it was the vector-intracellular fragment-extracellular fragment-vector. The recombinant expression vector pET-VPS-FC and the improved pET-VPS-SC of VP5 which was knocked out the gene fragment of transmembrane region were constructed. Then, the expression plasmid was transformed into BL21 （ DE3 ）. After IPTG induction, the recombinant protein was purified by Ni affinity chromatography and the gel filtration chromatography. [ Result ] The soluble expression IBDV VP5 was obtained. [ Conclusion ] The research laid the foundation for further studying the structure, and function of VP5 protein.

关 键 词：传染性法氏囊病病毒 VP5 跨膜区敲除 原核表达 

分 类 号：S188[农业科学—农业基础科学]