caa a22 4500 445822899 CHVBK 20180317145249.0 cr unu---uuuuu 170323e20110101xx s 000 0 eng 10.1007/s10616-010-9322-9 doi (NATIONALLICENCE)springer-10.1007/s10616-010-9322-9 Separation of supercoiled from open circular forms of plasmid DNA, and biological activity detection [Elektronische Daten] [Huangjin Li, Huaben Bo, Jinquan Wang, Hongwei Shao, Shulin Huang] To establish a cost-effective purification process for the large-scale production of plasmid DNA for gene therapy and DNA vaccination, a single anion-exchange chromatography (AEC) step was employed to purify supercoiled plasmid DNA (sc pDNA) from other isoforms and Escherichia coli impurities present in a clarified lysate. Two different size and conformation plasmids were used as model targets, and showed similar elution behavior in this chromatographic operation, in which sc pDNA was effectively separated from open circle plasmid DNA (oc pDNA) in a salt gradient. The process delivered high-purity pDNA of homogeneity of 95±1.1% and almost undetectable levels of endotoxins, genomic DNA, RNA and protein, at a yield of 65±8%. Furthermore, the transfection efficiency (29±0.4%) was significantly higher than that (20±0.1%) of a pDNA control. The present study confirms the possibility of using a single AEC step to purify sc pDNA from other isoforms and host contaminants present in a clarified E. coli lysate. Springer Science+Business Media B.V., 2010 Anion-exchange chromatography nationallicence Supercoiled plasmid DNA nationallicence Open circular plasmid DNA nationallicence Gene therapy nationallicence Li Huangjin School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut Bo Huaben School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut Wang Jinquan School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut Shao Hongwei School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut Huang Shulin School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut Cytotechnology Springer Netherlands 63/1(2011-01-01), 7-12 0920-9069 63:1<7 2011 63 10616 https://doi.org/10.1007/s10616-010-9322-9 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10616-010-9322-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Li Huangjin School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Bo Huaben School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Wang Jinquan School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Shao Hongwei School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Huang Shulin School of Life Science and Bio-Pharmaceutical, Guangdong Pharmaceutical University, 510006, Guangzhou, China aut NATIONALLICENCE 773 0- Cytotechnology Springer Netherlands 63/1(2011-01-01), 7-12 0920-9069 63:1<7 2011 63 10616 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer