利用绿色荧光蛋白GFP研究不同启动子在蝉棒束孢菌中的表达活性  

Green Fluorescent Protein Reported Expression Activities of Different Promoters in Isaria cicadae

作　　者：李栋 刘常利[2] 刘靖宇 孟俊龙[1,3] 王洪凯 LI Dong;LIU Changli;LIU Jingyu;MENG Junlong;WANG Hongkai(College of Food Science and Engineering,Shanxi Agricultural University,Taigu,Shanxi 030801,China;Biotechnology Institute,Zhejiang University,Hangzhou,Zhejiang 310058,China;Collaborative Innovation Center Quality and Efficience of Loess Plateau Edible Fungi in Shanxi,Taigu,Shanxi 030801,China)

机构地区：[1]山西农业大学食品科学与工程学院,山西太谷030801 [2]浙江大学生物技术研究所,浙江杭州310058 [3]黄土高原食用菌提质增效协同创新中心,山西太谷030801

出　　处：《食用菌学报》2020年第1期20-28,共9页Acta Edulis Fungi

基　　金：山西省煤基重点科技攻关项目(FT2014-03-01)。

摘　　要：利用农杆菌介导转化(Agrobacterium-mediated transformation,ATMT)的方法,将3个来源于蝉棒束孢菌(Isaria cicadae)、2个来源于稻瘟病菌(Magnaporthe grisea)的启动子在蝉棒束孢菌中驱动绿色荧光蛋白(green fluorescent protein,GFP)的表达。通过荧光显微镜观察GFP荧光亮度和荧光定量PCR检测GFP基因的转录水平,比较不同启动子的转录活性,并利用筛选出的高表达活性载体pKD5-GFP观察小G蛋白Rho在蝉棒束孢菌中的定位。结果表明:来源于稻瘟病菌的H3启动子在蝉棒束孢菌中对GFP基因具有较高的起始转录功能,小G蛋白Rho在孢子和新生菌丝阶段均有表达。研究结果可为在蝉棒束孢菌中进行外源基因表达和遗传研究提供参考。Using the Agrobacteriummediated transformation method and a green fluorescent protein(GFP)reporter gene,three promoters from Isaria cicadae and two promoters from Magnaporthe grisea were studied for expression activity in I.cicadae.GFP expressions driven by these promoters were compared in terms of fluorescence intensity by fluorescence microscopy and transcription level by fluorescence quantitative PCR.Then the vector with a high GFP expression level,pKD5-GFP,was selected and used to locate the small G protein Rho in I.cicadae.The results showed that the H3 promoter from M.grisea displayed high GFP transcription activity in I.cicadae,and Rho was expressed at both spore and new mycelium stages.These results provide a reference for genetic study and heterologous gene expression in I.cicadae.

关 键 词：蝉棒束孢菌 稻瘟病菌 小G蛋白 农杆菌介导转化 

分 类 号：S567.3[农业科学—中草药栽培]