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Updated: Jul 19, 2026

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures
Published on: January 5, 2011
Large-scale, nonchromatographic purification of plasmid DNA
Jason C Murphy1, Michael A Winters, Sangeetha L Sagar
1Vaccines and Biologics Research, Merck Research Laboratories, West Point, PA, USA.
Abstract:
A large-scale approach to the purification of plasmid DNA has been developed that overcomes many of the limitations of current chromatography-based processes. The process consists of a scaleable lysis using recombinant lysozyme and a rapid heating and cooling step followed by a selective precipitation with cetyltrimethylammonium bromide (CTAB). Calcium silicate batch adsorption is then utilized to remove residual genomic DNA, linear plasmid, open circular plasmid, endotoxin, detergents, and proteins. Finally, a concentration and diafiltration step utilizing ultrafiltration and a terminal sterile filtration complete the process. The final product exceeds the requirements for clinical-grade plasmid DNA, and the process has been scaled up to yield an average of 18 +/- 4 g (over five lots) of pharmaceutically pure plasmid DNA per 140 L of lysate (from approx 1.3 kg Escherichia coli dry cell weight).
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