双重PCR法同步检测菜豆晕疫病菌和萎蔫病菌

浙江农业学报

双重PCR法同步检测菜豆晕疫病菌和萎蔫病菌

（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水稻研究室，江苏连云港 222001；3中国检验检疫科学研究院植物检疫研究所，北京 100029）

出版日期:2015-09-25 发布日期:2015-09-23

Duplex PCR detection of Pseudomonas savastanoi pv. phaseolicola and Curtobacterium flaccumfaciens pv. flaccumfaciens in soybean#br#

（1 Lianyungang Entry\|Exit Inspection and Quarantine Bureau， Lianyungang 222042， China; 2 Institute of Rice， Lianyungang Academy of Agricultural Sciences， Lianyungang 222001， China; 3 Institute of Plant Quarantine， Chinese Academy of Inspection and Quarantine， Beijing 100029，China）

Online:2015-09-25 Published:2015-09-23

摘要/Abstract

摘要： 为建立同步检测菜豆晕疫病菌和细菌性萎蔫病菌的双重PCR方法，根据GenBank上公布的菜豆晕疫病菌argK基因特异性序列设计1对特异性引物PSPF1/PSPR2，将设计的引物与已发表的检测菜豆细菌性萎蔫病菌特异性引物CffF1/CffR2结合，经过条件优化，建立了双重PCR反应体系，并进行特异性和灵敏度验证。结果显示，采用双重PCR体系，能从菜豆晕疫病菌和细菌性萎蔫病菌基因组DNA以及人工模拟带菌大豆种子样品中扩增到特异性条带，表明本研究所建立的双重PCR检测方法能同时检测出菜豆晕疫病菌和细菌性萎蔫病菌，可用于口岸进口大豆中2种检疫性细菌的检测。

关键词: 菜豆晕疫病菌, 菜豆细菌性萎蔫病菌, 双重PCR, 检测方法

Abstract: The objective of this study was to establish a duplex PCR method for simultaneous detection of Pseudomonas savastanoi pv. phaseolicola（Psp） and Curtobacterium flaccumfaciens pv. flaccumfaciens（Cff）. Based on the argK gene of Psp in GenBank， the primers PSPF1/PSPR2 were designed. The duplex PCR assay was developed using the combining primers PSPF1/PSPR2 and CffF1/CffR2， which were specific primers for Cff gene. The reaction conditions were optimized and the specificity and sensitivity of the duplex PCR were tested. The expected DNA fragment was specifically amplified from the genomic DNA of Psp and Cff. Specificity was confirmed in the artificially inoculated soybean samples imported. Thus， the duplex PCR developed in this study could be used for the simultaneous detection of the two pathogens from imported soybean.

Key words: Pseudomonas savastanoi pv. phaseolicola, Curtobacterium flaccumfaciens pv. flaccumfaciens, duplex PCR, detection method

杨万风1，刘艳2，刘翔1，邵沛泽1，谌运清1，赵文军3. 双重PCR法同步检测菜豆晕疫病菌和萎蔫病菌[J]. 浙江农业学报.

YANG Wan\|feng1， LIU Yan2， LIU Xiang1， SHAO Pei\|ze1， CHEN Yun\|qing1， ZHAO Wen\|jun3. Duplex PCR detection of Pseudomonas savastanoi pv. phaseolicola and Curtobacterium flaccumfaciens pv. flaccumfaciens in soybean#br#[J]. .

[1]	杨万风, 刘艳, 陆辰晨, 邵沛泽, 谌运清, 赵文军. 双重PCR方法检测检疫性细菌洋葱腐烂病菌和菊基腐病菌[J]. 浙江农业学报, 2018, 30(3): 426-431.
[2]	杨万风1，刘艳2,刘翔1,邵沛泽1,谌运清1,赵文军3. 巢式PCR检测菜豆细菌性萎蔫病菌[J]. 浙江农业学报, 2015, 27(7): 1202-.