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Dye-ligand chromatography for the resolution and purification of restriction endonucleases

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Abstract

The resolution of restriction endonucleases from the same microorganism is conventionally achieved by lengthy fractionation protocols. We now report effective single-step procedures that exploit dye-ligand chromatography for the resolution and purification of restriction enzymes. After suitable initial screening, we demonstrated that resolution of two restriction activites can be achieved in one chromatographic step, and further purification can subsequently be effected using selected dye-adsorbents. Accordingly, we resolved in one step, Hpa I from Hpa II, Hind II from Hind III, and Sac I from Sac II. Furthermore, a three-step Chromatographic procedure has been developed to purify EcoRV suitable for commercial exploitation, as judged by the “overdigestion” and “cut-ligate-recut” quality control tests.

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Authors and Affiliations

  1. Applied Division, Research Center of Crete, Institute of Molecular Biology and Biotechnology, P.O. Box 1527, Iraklio, Crete, Greece

    G. Vlatakis, G. Skarpelis, I. Stratidaki & V. Bouriotis

  2. Institute of Biotechnology, University of Cambridge, Downing Street, CB2 3EF, Cambridge, UK

    Y. D. Clonis

Authors
  1. G. Vlatakis
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  2. G. Skarpelis
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  3. I. Stratidaki
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  4. V. Bouriotis
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  5. Y. D. Clonis
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Vlatakis, G., Skarpelis, G., Stratidaki, I. et al. Dye-ligand chromatography for the resolution and purification of restriction endonucleases. Appl Biochem Biotechnol 15, 201–212 (1987). https://doi.org/10.1007/BF02798449

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  • DOI: https://doi.org/10.1007/BF02798449

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