番茄溃疡病菌的环介导等温核酸扩增技术检测方法  

Detection of Clavibacter michiganensis subsp.Michiganensis by loop-mediated isothermal amplification

作　　者：封立平[1] 尼秀媚[1] 魏晓棠[1] 邵秀玲[1] 吴兴海[1] 厉艳[1] 

机构地区：[1]山东出入境检验检疫局,青岛266002

出　　处：《食品安全质量检测学报》2013年第4期1207-1212,共6页Journal of Food Safety and Quality

基　　金：质检总局科研项目(2009IK254)~~

摘　　要：目的建立番茄溃疡病菌的环介导等温核酸扩增技术检测方法。方法应用环介导等温核酸扩增技术(LAMP)对番茄溃疡病菌的16SrRNA特异性基因进行扩增,采用4对引物识别目的片段的6个特异性区域,62℃恒温1h完成扩增。结果设计的引物与对照菌皱纹假单胞菌、茄假单胞菌、丁香假单胞菌番茄致病变种都没有扩增反应,表现了较好的特异性。这些对照菌在茄科蔬菜上易引起类似症状。结论 LAMP检测程序便捷,所需设备简单,其结果可以通过肉眼观察来判断,比常规PCR灵敏且能更早得到反应结果,因此该技术具有更大的优势。Objective To establish a method for detection of Clavibacter michiganensis subsp. Michiganensis by loop-mediated isothermal amplification protocol (LAMP). Methods Detection of 16S rRNA of Clavibacter mi-chiganensis subsp. Michiganensis by a loop-mediated isothermal amplification protocol (LAMP) was evaluated in laboratory assays. LAMP amplified target DNA rapidly (1 h), isothermally (62℃) and with high-specificity based on four primers designed to recognize six independent sequences of target DNA. Results The LAMP primers did not react with suspensions of Pseudomonas corrugata, Pseudomonas solanacearum, and Pseudomonas syringae pv. to-mato. The specificity of this LAMP assay was higher than PCR. Conclusion This method has convenient testing procedures, the required equipment is simple, and the results can be judged by the naked eye. It is sensitive than con-ventional PCR reaction and the results can be obtained earlier, so it has a greater advantage of the technology.

关 键 词：番茄溃疡病菌 环介导等温核酸扩增技术 16SrRNA 

分 类 号：S436.412.1[农业科学—农业昆虫与害虫防治]