Reverse micellar extraction systems for the purification of pharmaceutical grade plasmid DNA
| Autor: | Carsten Voss, Nadine Streitner, Erwin Flaschel |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
Cell Extracts
Lysis two-phase systems Bioengineering Biology Applied Microbiology and Biotechnology Surface-Active Agents chemistry.chemical_compound Plasmid genetic vaccination Escherichia coli Technology Pharmaceutical reverse micelles Micelles Active ingredient Plasmid preparation Chromatography RNA DNA General Medicine Octanes gene therapy Molecular biology Quaternary Ammonium Compounds chemistry Agarose gel electrophoresis extraction Nucleic acid plasmid production Plasmids Biotechnology |
| Zdroj: | Journal of Biotechnology. 131:188-196 |
| ISSN: | 0168-1656 |
| DOI: | 10.1016/j.jbiotec.2007.06.011 |
| Popis: | Plasmid DNA as an active pharmaceutical ingredient (API) is gaining more and more importance. For the production of multigram quantities of this substance robust and scalable processes comprising several purification steps have to be designed. One main challenge is the initial separation of plasmid DNA and RNA in such a purification scheme. In this study we investigated the distribution of plasmid DNA and RNA in reverse micellar two-phase systems which is considered to be the basis for the development of an extractive purification step that can easily be integrated into common processes. For this purpose the distribution of the 4.6 kb plasmid pUT649 and Escherichia coli RNA in systems comprising isooctane, ethylhexanol, and the surfactant methyltrioctylammoniumchloride (TOMAC) under the influence of different salts was studied. Anion concentrations at which the partitioning behaviour for nucleic acids inverted (inversion point) were identified. Systems capable of separating RNA from plasmid DNA were further analysed and applied to extract RNA from plasmid DNA out of a preconditioned cleared lysate. The capability of reverse micellar systems for plasmid form separation was also shown by capillary and agarose gel electrophoresis. (C) 2007 Elsevier B.V. All rights reserved. |
| Databáze: | OpenAIRE |
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