DNA Extraction for Sensitive Detection of Shiga Toxin-Producing <i>Escherichia coli</i> in Food by Real-Time PCR Assays

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  • Mori Tetsuya
    Institute for Food and Environment Sciences Tokyo Kenbikyo-in Foundation
  • Nagao-Sato Sayaka
    Division of Microbiology, National Institute of Health Sciences
  • Kishino Kanae
    Institute for Food and Environment Sciences Tokyo Kenbikyo-in Foundation
  • Namba Toyohiko
    Institute for Food and Environment Sciences Tokyo Kenbikyo-in Foundation
  • Hara-Kudo Yukiko
    Division of Microbiology, National Institute of Health Sciences

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  • リアルタイムPCR法による腸管出血性大腸菌の食品での高感度検出のためのDNA抽出法
  • DNA Extraction for Sensitive Detection of Shiga Toxin-Producing Escherichia coli in Food by Real-Time PCR Assays

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Abstract

<p>Alkali-heat DNA extraction, a rapid and economical method, was evaluated for use in the detection of Shiga toxin-producing Escherichia coli in food using real-time PCR assays. Alkali-heat DNA extracts led to highly sensitive detection (102–104 CFU/mL) of stx and O-antigen genes in beef liver, ground beef, sliced pork, cheese, lettuce, radish sprouts, tomato, and spinach, equivalent to the sensitivity obtained using a commercial DNA extraction kit that utilizes proteinase K lysis, and silica membrane purification. Although there were differences in DNA concentration and purity between DNA extraction methods, the sensitivity of real-time PCR assays was similar. These results indicate that alkali-heat DNA extraction is a viable method when testing food products with real-time PCR assays for the presence of stx and O-antigen genes.</p>

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