Simultaneous molecular subtyping and shiga toxin gene detection in Escherichia coli using multiplex polymerase chain reaction

K L Hopkins; A. C. Hilton

doi:10.1046/j.1472-765x.2000.00695.x

Simultaneous molecular subtyping and shiga toxin gene detection in Escherichia coli using multiplex polymerase chain reaction

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Abstract

A robust random amplification of polymorphic DNA (RAPD)-polymerase chain reaction (PCR) protocol was developed for the combined epidemiological typing and shiga toxin detection of clinical shiga toxin-producing O157 and non-O157 Escherichia coli isolates. Using shiga toxin gene-specific primers, combined with two short 10-mer primers, in a multiplex shiga toxin/RAPD-PCR the fingerprints generated allowed differentiation between epidemiologically unrelated strains and allowed identification of a band amplified from the shiga toxin gene(s). Hybridization with a digoxigenin-labelled probe specific for stx1 and stx2 confirmed its identity. The combination of primers in this way allows valuable additional information to be gained from discriminatory RAPD profiles, with further benefits of time and cost savings over tests performed individually.

Original language	English
Pages (from-to)	122-125
Number of pages	4
Journal	Letters in Applied Microbiology
Volume	30
Issue number	2
DOIs	https://doi.org/10.1046/j.1472-765x.2000.00695.x
Publication status	Published - 1 Feb 2000

Bibliographical note

Funding: This work was funded by a grant provided by the Institute of Public and Environmental Health, University of Birmingham.

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10.1046/j.1472-765x.2000.00695.x

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title = "Simultaneous molecular subtyping and shiga toxin gene detection in Escherichia coli using multiplex polymerase chain reaction",

abstract = "A robust random amplification of polymorphic DNA (RAPD)-polymerase chain reaction (PCR) protocol was developed for the combined epidemiological typing and shiga toxin detection of clinical shiga toxin-producing O157 and non-O157 Escherichia coli isolates. Using shiga toxin gene-specific primers, combined with two short 10-mer primers, in a multiplex shiga toxin/RAPD-PCR the fingerprints generated allowed differentiation between epidemiologically unrelated strains and allowed identification of a band amplified from the shiga toxin gene(s). Hybridization with a digoxigenin-labelled probe specific for stx1 and stx2 confirmed its identity. The combination of primers in this way allows valuable additional information to be gained from discriminatory RAPD profiles, with further benefits of time and cost savings over tests performed individually.",

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AU - Hilton, A. C.

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AB - A robust random amplification of polymorphic DNA (RAPD)-polymerase chain reaction (PCR) protocol was developed for the combined epidemiological typing and shiga toxin detection of clinical shiga toxin-producing O157 and non-O157 Escherichia coli isolates. Using shiga toxin gene-specific primers, combined with two short 10-mer primers, in a multiplex shiga toxin/RAPD-PCR the fingerprints generated allowed differentiation between epidemiologically unrelated strains and allowed identification of a band amplified from the shiga toxin gene(s). Hybridization with a digoxigenin-labelled probe specific for stx1 and stx2 confirmed its identity. The combination of primers in this way allows valuable additional information to be gained from discriminatory RAPD profiles, with further benefits of time and cost savings over tests performed individually.

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Simultaneous molecular subtyping and shiga toxin gene detection in Escherichia coli using multiplex polymerase chain reaction

Abstract

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