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DNA Separation Mechanisms During Electrophoresis

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Capillary Electrophoresis of Nucleic Acids

Part of the book series: Methods in Molecular Biology ((MIMB,volume 162))

Abstract

This chapter describes the separation mechanisms used for DNA electrophoresis. The focus is on the concepts that may help the researcher understand the methodology, read the theoretical literature, analyze experimental data, identify the relevant separation regimes, and/or design optimization strategies. But first, let’s look at some key definitions. Since capillary electrophoresis (CE) is a “finish line” technique, the mobility μ(M) and the velocity v(M) of a molecule of size M (in bases or base pairs) in an electric field E are generally defined as: μ(M)= [v(M)]/E = L/[t(M)E] in which, L is the distance migrated during the elution time t(M). Clearly, this definition is valid only if v(M) is constant during the run. This requires time-independent and uniform (i.e., along the capillary) conditions (e.g., field, temperature, and so on), something that is rarely checked and is rather unlikely. This definition may thus lead, in some cases, to dubious conclusions (1). Successful separation of molecular sizes M1 and M2 requires the time spacing t1–t2 between these electrophoresis peaks to be larger than their full (time) width at half-maximum (FWHM), w1,2. A useful measure of the resolution is thus given by the separation factor S, which gives the smallest resolvable size difference: S = [(w1 + w2) × (M2 −M1)]/[2 × (t1 − t2)]

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Author information

Authors and Affiliations

  1. Department of Physics, University of Ottawa, Ottawa, Canada

    Gary W. Slater, Claude Desruisseaux & Sylvain J. Hubert

Authors
  1. Gary W. Slater
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  2. Claude Desruisseaux
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  3. Sylvain J. Hubert
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Editor information

Editors and Affiliations

  1. Australian Genome Research Facility, University of Queensland, Brisbane

    Keith R. Mitchelson

  2. Walter & Eliza Hall Institute, Parkville, Australia

    Keith R. Mitchelson

  3. Biochip Research and Development Center, State Key Laboratory for Biomembrane and Membrane Biotechnology, Tsinghua University, Beijing, China

    Jing Cheng

  4. Aviva Biosciences Corporation, San Diego, CA

    Jing Cheng

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© 2001 Humana Press Inc.

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Slater, G.W., Desruisseaux, C., Hubert, S.J. (2001). DNA Separation Mechanisms During Electrophoresis. In: Mitchelson, K.R., Cheng, J. (eds) Capillary Electrophoresis of Nucleic Acids. Methods in Molecular Biology, vol 162. Humana Press. https://doi.org/10.1385/1-59259-055-1:27

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  • DOI: https://doi.org/10.1385/1-59259-055-1:27

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-779-3

  • Online ISBN: 978-1-59259-055-1

  • eBook Packages: Springer Protocols

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