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Method for the characterization of size-exclusion chromatography media for preparative purification of DNA restriction fragments

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Biotechnology Techniques

Abstract

A protocol has been developed to characterize the potential of size exclusion chromatography media for the separation of DNA fragments at a preparative scale. A standard DNA mixture composed of λ-DNA cut with the restriction enzymes, AluI and HaeIII, was chromatographed on a column packed with the gel of interest. Fractions were collected and examined by PAGE. A digitized image of the gel was analyzed with the aid of a computer software program to determine the DNA fractionation range and selectivity curve for each chromatography gel. Four gels were characterized using this protocol. The effect of the elution buffer ionic strength on the fractionation of DNA was also investigated.

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Authors and Affiliations

  1. Mechanical and Industrial Engineering Department, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA

    Carlos M. Tellez

  2. Biotechnology Division, Bioprocess Engineering Group, 100 Bureau Dr. Stop 8312, National Institute of Standards and Technology, Gaithersburg, MD, 20899, USA

    Kenneth D. Cole

Authors
  1. Carlos M. Tellez
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  2. Kenneth D. Cole
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The US Government right to retain a non-exclusive royalty-freelicense in and to any copyright is acknowledged.

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Tellez, C.M., Cole, K.D. Method for the characterization of size-exclusion chromatography media for preparative purification of DNA restriction fragments. Biotechnology Techniques 13, 395–401 (1999). https://doi.org/10.1023/A:1008932407733

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  • DOI: https://doi.org/10.1023/A:1008932407733

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