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Contour-clamped homogeneous electric field electrophoresis of Staphylococcus aureus

Nature Protocols volume 1, pages 3028–3033 (2006)Cite this article

Abstract

Contour-clamped homogeneous electric field (CHEF) electrophoresis is a technique of pulsed-field gel electrophoresis that enables the resolution of large fragments of DNA that cannot be resolved by conventional gel electrophoresis. The procedure involves the application of controlled electric fields that change direction at a predetermined angle to samples of DNA that have been embedded in an agarose gel matrix and digested with a restriction endonuclease. Adjustment of the electrophoresis conditions enables the separation of DNA fragments with lengths from 10 kilobases up to 9 megabases in a size-dependent manner in agarose gels. The banding patterns can be used for epidemiological typing, the separated DNA can be immobilized onto a membrane and used for genetic mapping, or individual fragments can be extracted and used for downstream genetic manipulations. The protocol requires specialized equipment and can be completed in a maximum of 7 days.

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Figure 1: SmaI-digested chromosomal DNA of strains of Staphylococcus aureus following contour-clamped homogeneous electric field (CHEF) electrophoresis.

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Acknowledgements

This work was supported by the National Heath and Medical Research Council of Australia and the Western Australian Department of Health.

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Authors and Affiliations

  1. Gram-Positive Bacteria Typing and Research Unit, Curtin University of Technology, School of Biomedical Sciences, GPO Box U1987, Perth, Western Australia

    Frances G O'Brien & Warren B Grubb

  2. Department of Microbiology, Kuwait University, Box 24923, Safat, 13110, Kuwait

    Edet E Udo

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  1. Frances G O'Brien
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  2. Edet E Udo
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Correspondence to Frances G O'Brien or Edet E Udo.

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O'Brien, F., Udo, E. & Grubb, W. Contour-clamped homogeneous electric field electrophoresis of Staphylococcus aureus. Nat Protoc 1, 3028–3033 (2006). https://doi.org/10.1038/nprot.2006.382

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