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Proceedings Paper

Optimization of electric field strength for DNA sequencing in capillary gel electrophoresis
Author(s): John A. Luckey; Lloyd M. Smith
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Paper Abstract

Since its development, capillary gel electrophoresis has demonstrated the ability to separate DNA sequencing reactions at speeds roughly 25 times as great as conventional slab gel electrophoresis. These increased speeds are the result of using the more efficient dissipation of Joule heating by capillaries. However, to date there have been no studies which quantitate the advantages of disadvantages in operating these gels at high electric field strength. This work addresses this question by investigating the band-broadening of DNA sequencing reactions as they are separated through a fixed distance of gel at field strengths ranging from 50 V/cm to 400 V/cm. It is found that the bandwidths of DNA fragments do decrease with the higher field strengths due to a reduction in diffusional broadening. However, at sufficiently high electric field strengths, the bands begin to broaden again under the influence of an increasing thermal gradient across the diameter of the capillary. The result is an optimum electric field strength in the intermediate range of 100 - 250 V/cm depending on the length of fragments being separated. The relative importance of diffusion and thermal gradients are discussed and used to generate an equation that models the observed band broadening of DNA in capillary gel electrophoresis (CGE).

Paper Details

Date Published: 24 June 1993
PDF: 6 pages
Proc. SPIE 1891, Advances in DNA Sequencing Technology, (24 June 1993);
Show Author Affiliations
John A. Luckey, Univ. of Wisconsin/Madison (United States)
Lloyd M. Smith, Univ. of Wisconsin/Madison (United States)

Published in SPIE Proceedings Vol. 1891:
Advances in DNA Sequencing Technology
Richard A. Keller, Editor(s)

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