甘薯薯瘟病菌特异性PCR快速检测方法  

Specific and rapid PCR detection method of sweetpotato Ralstonia solanacearum

作　　者：张鸿[1] 刘中华[1] 林志坚[1] 许泳清[1] 李华伟[1] 王嘉滢 林赵淼[1] 李国良[1] 邱永祥[1] 纪荣昌[1] 罗文彬[1] 汤浩[1] 邱思鑫[1] Zhang Hong;Liu Zhonghua;Lin Zhijian;Xu Yongqing;Li Huawei;Wang Jiaying;Lin Zhaomiao;Li Guoliang;Qiu Yongxiang;Ji Rongchang;Luo Wenbin;Tang Hao;Qiu Sixin(Institute of Crop Sciences,Fujian Academy of Agricultural Sciences Scientific Observing&Experimental Station of Tuber&Root Crops in South China,Ministry of Agriculture Technical Research Center of Specialty Dry Crop Variety Breeding of Fujian,Fuzhou 350013,Fujian,China;Jinshan College,Fujian Agriculture&Forestry University,Fuzhou 350002,Fujian,China)

机构地区：[1]福建省农业科学院作物研究所/农业部南方薯类科学观测实验站/福建省特色旱作物品种选育工程技术研究中心,福建福州350013 [2]福建农林大学金山学院,福建福州350002

出　　处：《江苏师范大学学报（自然科学版）》2019年第4期37-41,共5页Journal of Jiangsu Normal University：Natural Science Edition

基　　金：省属公益类科研院所基本科研专项(2017R1026-4);现代农业产业技术体系细菌病防控项目(CARS-10-B14);福建省农业科学院创新团队项目(STIT2017-2-3);福建省农业科学院自由探索科技创新项目(ZYTS2019007)

摘　　要：针对目前甘薯薯瘟病鉴定耗时长,且缺乏特异性标记的问题,首先根据薯瘟病菌Ralstonia solanacearum特异性基因orf428设计了一对特异性分子检测引物,对13株薯瘟病菌、3株甘薯其他病原细菌和12株其他7种寄主青枯菌进行PCR检测,结果显示,只有薯瘟病菌能特异扩增1641 bp的目标序列,且对菌体DNA最低检出率达到1 pg/μL.其次,通过水煮法快速提取DNA,简化了DNA提取方法;在PCR体系中加入体积分数为1%的二甲基亚砜(DMSO)后,扩增条带更为明亮,优化了PCR检测体系.此技术易于操作、灵敏度高、特异性强,可用于对病残体、发病植株和种苗中的薯瘟病菌检测,以对甘薯薯瘟病进行预测、监测及防治具有重要意义.For identification of sweetpotato bacterial wilt caused by Ralstonia solanacearum,traditional method is time consuming and lack of specific markers,a new specific and rapid PCR detection method was reported.The designed two PCR primers were based on the specific gene orf428 in sweetpotato Ralstonia solanacearum.The PCR test results showed that the target 1641 bp sequence could only be amplified in sweetpotato Ralstonia solanacearum,compared to 12 Ralstonia solanacearum strains from other 7 hosts and other 3 pathogenic bacteria strains in sweetpotato.The lowest amount of DNA needed for this detection was 1 pg/μL.The bacterial DNA extraction method was simplified by using the boiled leaching solution of disease samples for PCR amplification directly.The PCR reaction system was also optimized,which is adding 1%volume fraction DMSO to make the electrophoretic band brighter.The easy operated,highly specific,accurate and sensitive method can be further applied in forecast and long-term monitering of sweetpotato bacterial wilt,which is of great significance to take appropriate measures to control the disease.

关 键 词：甘薯薯瘟病 青枯菌 PCR 检测 

分 类 号：S531[农业科学—作物学] S432.1