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Molecular Pathology

DNA Purification

Anion-Exchange Chromatography


Author: Rodney E. Shackelford, D.O., Ph.D. (see Reviewers page)
Revised: 14 April 2012, last major update April 2012
Copyright: (c) 2008-2011, PathologyOutlines.com, Inc.

Definition
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● A subtype of ion exchange chromatography (Wikipedia)
● A rapid and efficient method of nucleic acid purification which removes lipids, proteins and other cellular constituents in one rapid step
● Uses a positively charged solid resin column that captures negatively charged nucleic acids
● Prior to use, the column is equilibrated with a buffer containing anions and the crude nucleic acid preparation (usually a cell lysate) is added to the column
● Negatively charged nucleic acids are preferentially retained in the column while other cellular constituents are washed out, allowing one-step purification
● After several washes to remove unwanted cellular constituents, the nucleic acids are removed with a high-salt, low pH wash
● Commonly, the elution profiles of RNA and DNA overlap and other steps must be undertaken if pure RNA or DNA is required
● Diethylaminoethyl (DEAE) covalently linked to sepharose (a polysaccharide polymer) is used in anion-exchange chromatography

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