| Using a single hydrophobic-interaction chromatography to purify pharmaceutical-grade supercoiled plasmid DNA from other isoforms. | |
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MedLine Citation:
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PMID: 23013372 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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Context: The recent developments in non-viral gene therapy and DNA vaccine have fostered the development of efficient plasmid DNA (pDNA) purification processes. Objectives: This work aimed to establish a cost-effective purification process for the large-scale production of plasmid DNA for gene therapy and DNA vaccine. Materials and methods: E. coli DH5α harboring pCDNA3.1-GFP (7200 base pairs) was used as a model plasmid. Hydrophobic-interaction chromatography (HIC) was employed to purify supercoiled plasmid DNA (sc pDNA). Results: With this method, not only host contaminants, but also open circular plasmid DNA (oc pDNA) could be removed from sc pDNA. Anion-exchange HPLC analysis proved that the recovery of HIC could reach 75%. The plasmid DNA exhibited high purity with supercoiled percentage of 98 ± 1.2% and undetectable residual endotoxins, genomic DNA, RNA and protein. The purity of pDNA had nothing to do with the flow rate in the range at least up to 400 cm/h. Liposomes transfection experiment prove that the purified pDNA in this article had higher transfection efficiency than the control pDNA. Discussion and conclusion: In the present work, we confirmed the possibility of separation of sc pDNA from oc pDNA and other host contaminants using a single HIC chromatography. |
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Authors:
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Huaben Bo; Jinquan Wang; Qizhu Chen; Han Shen; Fenglin Wu; Hongwei Shao; Shulin Huang |
Publication Detail:
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Type: JOURNAL ARTICLE Date: 2012-9-27 |
Journal Detail:
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Title: Pharmaceutical biology Volume: - ISSN: 1744-5116 ISO Abbreviation: Pharm Biol Publication Date: 2012 Sep |
Date Detail:
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Created Date: 2012-9-27 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 9812552 Medline TA: Pharm Biol Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
Affiliation:
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Institute of Genetic Engineering, Southern Medical University , Guangzhou , PR China. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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