Effect of alcohol on purification of plasmid DNA using ion-exchange membrane

Abstract

An ion-exchange membrane, positively charged with quaternary ammonium groups on polyethersulfone copolymer, was used to purify plasmid DNA from bacterial lysate in this work. Batch operation was used to examine how plasmid purification was affected after the incorporation of alcohols or a chaotropic salt, guanidine hydrochloride, into the washing buffers. The incorporation of alcohol had two opposite effects on the binding strengths between nucleic acids and the partially hydrophobic membrane. The binding can be enhanced by the reduced dielectric constant and on the other hand be decreased by the increased hydrophobicity. At alcohol concentration above 40% (v/v), enhancement of plasmid yield was observed for the use of methanol, ethanol, or isopropanol. The use of guanidine hydrochloride by itself decreased the plasmid yield. Such decreases, however, could be restored when guanidine hydrochloride was used together with alcohol. When membrane chromatography was used to purify plasmid using the washing buffer which contained 40% isopropanol and 1.5 M NaCl, protein and RNA were undetected in the eluted plasmid. (C) 2004 Elsevier B.V. All rights reserved.