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Comparison of DNA Extraction Methods for the Detection of Foodborne Pathogenic Bacteria from Livestock Manure Composts

퇴비에서 식중독균 검출을 위한 DNA 추출 방법 비교

  • Kim, Sung-Youn (Division of Safety Analysis, Experiment & Research Institute National Agricultural Products Quality Management Service) ;
  • Seo, Dong-Yeon (Division of Safety Analysis, Experiment & Research Institute National Agricultural Products Quality Management Service) ;
  • Moon, Ji-Young (Division of Safety Analysis, Experiment & Research Institute National Agricultural Products Quality Management Service)
  • 김성연 (국립농산물품질관리원 시험연구소 안전성분석과) ;
  • 서동연 (국립농산물품질관리원 시험연구소 안전성분석과) ;
  • 문지영 (국립농산물품질관리원 시험연구소 안전성분석과)
  • Received : 2019.07.10
  • Accepted : 2019.11.28
  • Published : 2019.12.30

Abstract

This study investigated the efficacy of DNA extraction methods for real-time PCR detection of foodborne pathogenic bacteria in livestock manure composts. Livestock manure composts were inoculated with Escherichia coli O157:H7, Salmonella Typhimurium, Listeria monocytogenes, Bacillus cereus and incubated in enrichment broth. For DNA extraction, enriched samples were treated following boiling method, by chloroform, C18 powder, and proteinase K. As a result, 4 species of bacteria were detected by real-time PCR when subjected to boiling for 30 min and treated with proteinase K. These results suggest that detection of foodborne pathogens by real-time PCR from livestock manure composts could be applicable using effective DNA extraction methodology such as the boiling method or proteinase K.

본 연구에서는 가축분퇴비에 존재할 수 있는 식중독균의 검출을 위하여 기존의 배양을 이용한 방법을 대체할 수 있는 real-time PCR을 적용하고자 하였으며, 이에 따라 유전자 증폭에 영향을 미치는 DNA 추출 방법에 따른 식중독균 검출 효율을 비교하였다. 적용한 방법은 가열 처리, 유기용매 및 흡착제 처리, 효소 처리의 3가지로 구분할 수 있으며, 각 방법에 따른 DNA의 검출 효율을 실험 결과로 나타내었다. 가열 처리 방법에서는 가열 시간의 증가에 따라 DNA 검출 효율이 높아지는 경향을 나타냈으며, 유기용매 및 흡착제는 효과를 나타내지 않았고, 효소 처리의 경우에는 그람 양성균 보다는 그람 음성균의 DNA가 추출 효율이 더 높은 것으로 나타났다. 결론적으로 퇴비에서 30분 이상의 가열 처리와 효소의 처리를 통한 DNA 추출 방법은 real-time PCR을 적용한 식중독균 검출에 적합한 것으로 판단된다.

Keywords

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