Document Detail

Using a single hydrophobic-interaction chromatography to purify pharmaceutical-grade supercoiled plasmid DNA from other isoforms.
MedLine Citation:
PMID:  23013372     Owner:  NLM     Status:  Publisher    
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.
Huaben Bo; Jinquan Wang; Qizhu Chen; Han Shen; Fenglin Wu; Hongwei Shao; Shulin Huang
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-9-27
Journal Detail:
Title:  Pharmaceutical biology     Volume:  -     ISSN:  1744-5116     ISO Abbreviation:  Pharm Biol     Publication Date:  2012 Sep 
Date Detail:
Created Date:  2012-9-27     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9812552     Medline TA:  Pharm Biol     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Institute of Genetic Engineering, Southern Medical University , Guangzhou , PR China.
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