Purification of supercoiled plasmid DNA using chromatographic processes |
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Authors: | Ferreira G N Cabral J M Prazeres D M |
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Institution: | Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Lisboa, Portugal. |
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Abstract: | The interest in purifying injectable-grade plasmid DNA has increased with the development of gene therapy and DNA vaccination technologies. In this paper we develop a method for purifying a 4.8 kb plasmid based on chromatographic processes. An NaCl gradient was optimized on a Q Sepharose column and plasmid was eluted at 800-820 mM NaCl in a broad peak. Supercoiled plasmid was isolated after a final Sepharcryl S1000 SF gel filtration step. Final plasmid preparation was depleted of proteins and RNA, as revealed by the BCA assay and 1% agarose gel electrophoresis. |
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