Separating plasmids from contaminants using hydrophobic or hydrophobic and ion exchange chromatography

Abstract

Long chain nucleic acids such as plasmids are separated from contaminants including protein, RNA, DNA and mixtures thereof by using hydrophobic interaction chromatography or hydrophobic interaction chromatography and ion exchange chromatography. To carry out hydrophobic interaction chromatography, first and second columns containing first and second column materials are used. The first column material adsorbs protein and RNA at a salt concentration at which plasmid is not adsorbed to produce an eluate containing plasmid and DNA. The second column material adsorbs plasmid and DNA from the eluate at a salt concentration which the first column material adsorbs protein and RNA but not the plasmid. The plasmid is eluted from the second column material at a salt concentration at which plasmid is eluted. A tandem column may be formed by connecting in series the first and second columns. The connection between the columns is broken before eluting plasmid from the second column material. Ion exchange chromatography is carried out in addition to the hydrophobic interaction chromatography by adsorbing plasmid from an eluate from the second column material onto an ion exchange chromatography material, and eluting the adsorbed plasmid.