油菜菌核病菌激发子基因SsXYN克隆及表达载体构建  

Cloning of Sclerotinia sclerotiorum elicitor gene SsXYN and the expression vector construction

作　　者：蒋冰心[1] 吴群[1] 高智谋[1] 张华建[1] 

机构地区：[1]安徽农业大学植物保护学院,合肥230036

出　　处：《安徽农业大学学报》2013年第4期631-635,共5页Journal of Anhui Agricultural University

基　　金：国家自然科学青年基金项目(31101426);公益性行业(农业)科研专项(201103016)共同资助

摘　　要：以核盘菌菌株NGA4提取的总RNA为模版,利用RT-PCR技术扩增获得SsXYN基因的全长cDNA片段,将其克隆到pMD19-T载体上,菌落PCR、酶切鉴定和cDNA测序结果表明成功克隆了核盘菌SsXYN基因。从pMD19-T:SsXYN载体上,用BamH I和Sal I双酶切切下目的基因片段,将该基因片段连接到原核表达载体pET32a中。菌落PCR和酶切鉴定结果显示成功构建了表达载体pET32a:SsXYN。利用热击法将该重组表达载体导入大肠杆菌BL21,获得携带SsXYN基因的大肠杆菌菌株,为进一步研究激发子诱发植物抗病性机理奠定了基础。Total RNA was extracted from Sclerotinia sclerotiorum NGA4mycelia and used as the template to amplify the full length of cDNA of SsXYN gene by RT-PCR,and the gene fragment was subsequently cloned into pMD19-T vector.The results of bacterial colony PCR,enzyme analysis and sequencing analysis indicated that the S.sclerotiorum SsXYN gene was successfully cloned.SsXYN gene was digested completely from pMD19-T ： SsXYN by BamH I and Sal I,and then was cloned into expression vector pET32a.The results of bacterial colony PCR and enzyme analysis showed the expression vector pET32a ： SsXYN was successfully constructed.After that,the recombinant expression vector was carried into E.coli BL21 by heat-shock transformation,and the strain of E.coli BL21 carrying SsXYN gene was obtained.The study provides a basis for our understanding of the elicitor SsXYN downstream signaling pathway in plant immunity.

关 键 词：核盘菌 SsXYN 激发子 

分 类 号：S435.654[农业科学—农业昆虫与害虫防治]