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Toxins 2015, 7(11), 4745-4757; doi:10.3390/toxins7114745

A Simple and Rapid Procedure for the Detection of Genes Encoding Shiga Toxins and Other Specific DNA Sequences

1
Depratment of Molecular Biology, University of Gdańsk, Wita Stwosza 59, 80-308 Gdańsk, Poland
2
Department of Bacteriology, National Institute of Public Health-Public Institute of Hygiene, 24 Chocimska Street, 00-791 Warsaw, Poland
3
Laboratory of Molecular Biology (affiliated with the University of Gdansk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Wita Stwosza 59, 80-308 Gdansk, Poland
*
Author to whom correspondence should be addressed.
Academic Editor: Vernon L. Tesh
Received: 28 September 2015 / Revised: 26 October 2015 / Accepted: 10 November 2015 / Published: 13 November 2015
(This article belongs to the Section Bacterial Toxins)
View Full-Text   |   Download PDF [1111 KB, uploaded 13 November 2015]   |  

Abstract

A novel procedure for the detection of specific DNA sequences has been developed. This procedure is based on the already known method employing PCR with appropriate primers and a sequence-specific DNA probe labeled with the fluorescent agent 6-carboxylfluorescein (FAM) at the 5′ end and the fluorescence quencher BHQ-1 (black hole quencher) at the 3′ end. However, instead of the detection of the fluorescence signal with the use of real-time PCR cyclers, fluorescence/luminescence spectrometers or fluorescence polarization readers, as in all previously-reported procedures, we propose visual observation of the fluorescence under UV light directly in the reaction tube. An example for the specific detection of the Shiga toxin-producing Escherichia coli (STEC) strains, by detecting Shiga toxin genes, is demonstrated. This method appears to be specific, simple, rapid and cost effective. It may be suitable for use in research laboratories, as well as in diagnostic units of medical institutions, even those equipped only with a thermocycler and a UV transilluminator, particularly if rapid identification of a pathogen is required. View Full-Text
Keywords: DNA amplification; detection methods; PCR product detection; shiga toxin-producing Escherichia coli; stx genes DNA amplification; detection methods; PCR product detection; shiga toxin-producing Escherichia coli; stx genes
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Nejman-Faleńczyk, B.; Bloch, S.; Januszkiewicz, A.; Węgrzyn, A.; Węgrzyn, G. A Simple and Rapid Procedure for the Detection of Genes Encoding Shiga Toxins and Other Specific DNA Sequences. Toxins 2015, 7, 4745-4757.

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