PuGV-Ps增效蛋白质基因特征及其整合载体的构建  

Characteristics of enhancin gene from Pseudaletia unipuncta granulovirusPs and construction of its integration vector

作　　者：韩光杰[1] 赵松[2] 刘琴[1] 李传明[1] 徐健[1] 

机构地区：[1]江苏里下河地区农业科学研究所,江苏扬州225007 [2]江苏省血吸虫病防治研究所,江苏无锡214064

出　　处：《江苏农业学报》2015年第2期279-284,共6页Jiangsu Journal of Agricultural Sciences

基　　金：公益性行业(农业)科研专项(201103021);江苏省农业科技自主创新基金项目[CX(13)5069];江苏省自然科学基金项目(BK20141283);江苏省农业科技支撑计划项目(BE2014361);江苏省农业三新工程项目[SXGC(2013)256];扬州市农业科技攻关项目(2012095)

摘　　要：为了明确增效蛋白质基因特性,获得重组增效苏云金杆菌,本试验通过生物信息学方法分析了转宿主东方粘虫颗粒体病毒（Pu GV-Ps）增效蛋白质基因（En）的特征及二级结构,利用In-fusion技术构建了整合载体。基于Pu GV-Ps增效蛋白质基因进化树显示,颗粒体病毒与多角体病毒为2个明显的分支,美洲粘虫颗粒体病毒（Pu GV）与转宿主东方粘虫颗粒体病毒的增效蛋白质基因遗传距离极小。6种颗粒体病毒增效蛋白质氨基酸序列比对结果显示,增效蛋白质N端相似性较高,均含有锌离子结构域（244位）与催化域（526～565位）;增效蛋白质基因二级结构中α螺旋和无规则卷曲占整个片段的73%,且N端以不规则卷曲为主;利用改进的In-fusion技术构建整合载体,双酶切结果显示增效蛋白质基因连接方向正确,p LTV1-En构建成功;通过高温整合获得重组工程菌Bt71En。表明,Pu GV-Ps与Pu GV增效蛋白质基因同源性较高,其二级结构复杂,影响载体构建,通过高温处理连接片段可以明显提高载体的连接效率。The characteristics of enhancin gene from Pseudaletia unipuncta granulovirus-Ps （PuGV-Ps） and the influ- ence of secondary structure on construction of integration vector were analyzed by bioinformatics in this study. The granulosis virus and polyhedrosis virus were grouped into two clades based on the structure of enhancin gene, and there was an extremely short genetic distance of enhancin between PuGV-Ps and Pseudaletia unipuncta granulovirus（ PuGV）, suggestive a high simi- larity. N-terminal of enhancin gene from different granuloviruses were highly similar, all including zinc-binding domain （244） and catalytic domain （526-565）. The alpha helix and random coil in the complete enhancin gene accounted for 73% of the secondary structure, and the random coil dominated N-terminal. The integration vector of pLTV1-En successfully constructed by improved technology of In-fusion was used to produce recombinant Bt71En, and the connection efficiency could be improved by high temperature.

关 键 词：转宿主粘虫颗粒体病毒 增效蛋白质 整合载体 

分 类 号：S182[农业科学—农业基础科学]