The Korean Journal of Mycology (한국균학회지)

The Korean Society of Mycology (한국균학회)
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Development of a Loop-mediated Isothermal Amplification Detection Assay for Verticillium dahliae Infection in Chrysanthemum

국화에 발생하는 반쪽시들음병균 Verticillium dahliae 검출용 등온 증폭법 개발

  • Back, Chang-Gi (Horticultural and Herbal Crop Environment Division, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Park, Mi-Jeong (Horticultural and Herbal Crop Environment Division, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Han, Kyung-Sook (Planning and Coordination Division, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Park, Jong-Han (Horticultural and Herbal Crop Environment Division, National Institute of Horticultural and Herbal Science, Rural Development Administration)
  • 백창기 (국립원예특작과학원 원예특작환경과) ;
  • 박미정 (국립원예특작과학원 원예특작환경과) ;
  • 한경숙 (국립원예특작과학원 기획조정과) ;
  • 박종한 (국립원예특작과학원 원예특작환경과)
  • Received : 2019.12.19
  • Accepted : 2019.12.20
  • Published : 2019.12.31
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Abstract

Verticillium wilt disease is caused by a fungal plant pathogen Verticillium dahliae, which attacks commercial crops such as chrysanthemum. The conventional methods so far used to identify this fungal pathogen require high expertise and are time-consuming. Therefore, in this study, we developed an assay for the rapid and specific detection of V. dahliae infection using loop-mediated isothermal amplification (LAMP) method. For this assay, four primers for LAMP were designed for targeting cellulose-growth-specific protein partial mRNA gene in Verticillium dahliae. Under standard condition, the optimum reaction temperature for amplification is around 60 ℃ within 60 minutes. This LAMP assay was designed to amplify only present in V. dahliae. When this LAMP assay applied to the DNAs for four other soil-borne fungi and host plants, no amplification was detected. Therefore, this LAMP assay we developed for V. dahliae is expected to do detection at the early stage of its infection. The fast and reliable detection method will allow us to develop effective management system to monitor and control infection of this pathogen in chrysanthemum plant.

국화에 발생하는 반쪽시들음병은 Veriticillium dahliae에 의해 발생하는 진균병으로 국화 재배농가에 상당한 경제적 손실을 야기한다. 일반 식물병원균을 동정하는 방법으로는 병원균을 진단하기까지 상당한 시간이 소요된다. 본 연구에서는V. dahliae를 신속하고 특이적으로 진단하기 위하여 등온증폭기술 (Loop-mediated isothermal amplification, LAMP)을 적용한 검출법을 개발하였다. 이 방법은 반쪽시들음병균의 cellulose-growth-specific protein partial mRNA 유전자 염기서열을 이용하여 4개의 특이 프라이머 세트를 제작하였다. 최적 반응조건 및 시간은 60℃ 내외의 온도조건에서 60분 이내에서 가장 효율이 좋은 것으로 나타났다. 이 등온증폭 검출법은 4종의 토양전염성 병원균과 기주식물의 DNA에는 반응하지 않았다. 따라서 반쪽시들음병균 등온증폭법을 활용한다면 병원균의 감염 유무를 조기에 신속하게 진단할 수 있고, 반쪽시들음병을 효율적으로 모니터링하고 방제할 수 있을 것으로 기대한다.

Keywords

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