菜豆晕疫病菌环介导等温核酸扩增检测方法的建立  

Development of loop-mediated isothermal amplification(LAMP) method for detection of Pseudomonas savastanoi pv. phaseolicola

作　　者：杨万风[1] 刘艳[2] 刘翔[1] 邵沛泽[1] 谌运清[1] 赵文军[3] 

机构地区：[1]连云港出入境检验检疫局,江苏连云港222042 [2]连云港市农业科学院,江苏连云港222001 [3]中国检验检疫科学研究院,北京100029

出　　处：《江苏农业学报》2014年第6期1321-1327,共7页Jiangsu Journal of Agricultural Sciences

基　　金：国家质检总局科技计划项目(2013IK277)

摘　　要：为了建立菜豆晕疫病菌的环介导等温扩增方法,本试验利用菜豆晕疫病菌arg K特异性序列设计环介导等温核酸扩增(LAMP)引物,进行反应条件和反应体系的优化,并进行特异性和灵敏度验证。特异性检测结果显示,5株菜豆晕疫病菌的LAMP产物呈阳性结果,而参试的其他病原菌不产生扩增反应。LAMP检测基因组DNA和菌悬液时,其灵敏度分别达到0.632×10-4ng/μl和7.3×102CFU/ml,该方法比常规PCR灵敏度高100倍。表明本研究所建立的LAMP检测方法简便、快速、准确、灵敏,可有效应用于口岸菜豆晕疫病菌的检测。To develop a loop-mediated isothermal amplification（LAMP） method for detecting Pseudomonas savastanoi pv.phaseolicola,LAMP primers were designed based on the arg K gene of P.savastanoi pv.phaseolicola,the reaction conditions were optimized,and the specificity and sensitivity of LAMP were tested.The amplification results revealed that five isolates of P.savastanoi pv.phaseolicola were positive,while other pathogens were negative.LAMP method showed sensitivities of 6.32×10^-5ng/μl and 7.3×10^2CFU/ml when detecting genomic DNA and bacterial suspension of P.savastanoi,100 times higher than conventional PCR assay.The LAMP method developed in this study was simple,fast,specific and sensitive,for detecting P.savastanoi pv.phaseolicola in port quarantine.

关 键 词：菜豆晕疫病菌 环介导等温扩增 检测 

分 类 号：S432.1[农业科学—植物病理学]