MAO Fu-rong,LI Fei-wu,LIU Yan-ni,et al.Qualitative PCR Detection Method for Clavibacter michiganensis subsp.Michiganensis[J].Northern Horticulture,2015,39(05):128-131.[doi:10.11937/bfyy.201505037]
番茄细菌性溃疡病菌的定性PCR检测方法
- Title:
- Qualitative PCR Detection Method for Clavibacter michiganensis subsp.Michiganensis
- 文献标志码:
- A
- 摘要:
- 以2个番茄溃疡病菌株和其它6种植物病原菌为试验材料，以micA、cytC、TomA等3个特异性基因为检测靶标，采用检测引物设计与优化、特异性测试、灵敏度测试等方法，研究建立番茄溃疡病菌的定性PCR检测方法。结果表明：依据这3个基因建立的PCR检测方法可特异、精准检测番茄溃疡病菌，其中，以TomA基因为检测靶标的方法灵敏度可达到5 copy/μL或103 cfu/mL，优于另外2种方法，为番茄溃疡病菌的早期诊断提供了快速、准确的技术手段。
- Abstract:
- Using two Clavibacter michiganensis subsp.michiganensis strains and six kinds of plant pathogen as experimental materials，through primers design，specificity and sensitivity testing，a PCR method for detection of Clavibacter michiganensis subsp.michiganensis by employing three specific genes of micA，cytC，and TomA were established.The results showed that this developed PCR method resulted in a specific and accurate detection of Clavibacter michiganensis subsp.michiganensis.In particular，the sensitivity of the detection method using TomA gene as test target could reach 5 copy/μL or 103 cfu/mL，which was more sensitive than the other two methods by employing micA and cytC gene，respectively.This study provided a fast and precise technological means for the early diagnosis of Clavibacter michiganensis subsp.michiganensis.
参考文献/References:
[1] Wittmann J，Gartemann K H，Eichenlaub R，et al.Genomic and molecular analysis of phage CMP1 from Clavibacter michiganensis subspecies michiganensis[J].Bacteriophage，2011，1(1)：6-14. [2] 罗来鑫，赵廷昌，李建强，等.番茄细菌性溃疡病研究进展[J].中国农业科学，2004，37(8):1144-1150. [3] 赵文军，夏明星，陈红运，等.番茄溃疡病菌PCR快速检测技术[J].植物检疫，2007，21(2):75-77. [4] 王蕊，罗来鑫，李健强.不同来源番茄溃疡病菌致病力差异研究[J].植物保护，2010，36(1):73-77. [5] 李江利，欧阳林杰，郭淼淼，等.河南省番茄溃疡病的发现与分子确诊[J].河南农业大学学报，2011，45(3):349-352. [6] 王念武，王婷，沈建国，等.基于锁式探针的番茄溃疡病菌实时荧光PCR快速检测[J].中国农业科学，2014，47(5):903-911. [7] 闵现华，刘菁，韩跃武，等.Nested-PCR检测种传番茄细菌性溃疡病菌[J].西北农业学报，2011，20(7):28-31. [8] 闵现华，韩跃武，刘菁，等.种传番茄溃疡病菌直接PCR和免疫捕捉PCR检测方法之比较[J].植物检疫，2010，24(4):12-16. [9] 封立平，尼秀媚，魏晓棠，等.番茄溃疡病菌的环介导等温核酸扩增技术检测方法[J].食品安全质量检测学报，2013，4(4):1207-1212. [10]Yasuhara-Bell J，Kubota R，Jenkins D M，et al.Loop-mediated amplification of the Clavibacter michiganensis subsp. michiganensis micA gene is highly specific[J].Phytopathology，2013，103(12)：1220-1226. [11]Cho M S，Lee J H，Her N H，et al.A quantitative and direct PCR assay for the subspecies-specific detection of Clavibacter michiganensis subsp.michiganensis based on a ferredoxin reductase gene[J].Journal of Microbiology，2012，50(3)：496-501. [12]Gartemann K H，Abt B，Bekel T，et al.The genome sequence of the tomato-pathogenic actinomycete Clavibacter michiganensis subsp. michiganensis NCPPB382 reveals a large island involved in pathogenicity[J].Journal of Bacteriology，2008，190(6)：2138-2149. [13]王念武，王婷，沈建国，等.超分支滚环扩增技术快速检测番茄溃疡病菌[J].植物检疫，2013，27(5):66-70. [14]Bach H J，Jessen I，Schloter M，et al.A TaqMan-PCR protocol for quantification and differentiation of the phytopathogenic Clavibacter michiganensis subsp.sepedonicus[J].Journal of Microbiological Methods，2003，52(1)：85-91. [15]Holtsmark I，Mantzilas D，Eijsink V G，et al.Purification，characterization，and gene sequence of michiganin A，an actagardine-like lantibiotic produced by the tomato pathogen Clavibacter michiganensis subsp.michiganensis[J].Applied and Environmental Microbiology，2006，72(9)：5814-5821. [16]吴兴海，邵秀玲，厉艳，等.应用分子生物学方法快速检测番茄溃疡病菌的研究[J].江西农业学报，2006，18(2):5-7. [17]付鹏，郭亚辉，张晓梅，等.番茄溃疡病菌分子检测技术[J].江苏农业学报，2005，21(2):118-122.
相似文献/References:
[1]顾志光,徐文凤,范玲超,等.拮抗放线菌固态发酵菌剂及其盆栽促生效果[J].北方园艺,2017,41(08):124.[doi:10.11937/bfyy.201708028]
GU Zhiguang,XU Wenfeng,FAN Lingchao,et al.Solid Fermentation of Antagonistic Actinomycetes and Effect of Promoting Plant Growth in Pot[J].Northern Horticulture,2017,41(05):124.[doi:10.11937/bfyy.201708028]
[2]高娃,张冬梅,高振江,等.番茄溃疡病病原的鉴定和抗性品种的筛选[J].北方园艺,2019,43(23):20.[doi:10.11937/bfyy.20191132]
GAO Wa,ZHANG Dongmei,GAO Zhenjiang,et al.Identification of Pathogeny of Tomato Canker Disease and Screening of Resistance Varieties[J].Northern Horticulture,2019,43(05):20.[doi:10.11937/bfyy.20191132]
备注/Memo
第一作者简介：毛芙蓉（1980-），女，本科，助理研究员，研究方向为蔬菜病虫害综合防治。E-mail：lotusfurmao@gmail.com. 基金项目：吉林省重点科技攻关资助项目（20140204032NY)。