Separation and Size Assessment of dsDNA Fragments by Anion-Exchange Chromatography (AEX)

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Hua Yang, Stephan M. Koza, Weibin Chen
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Anion-exchange chromatography is robust, reproducible, yields quantitative information, and is easy to be automated. We demonstrate here that a 1 kb Plus DNA Ladder, ranging from 0.1 kbp to 10.0 kbp dsDNA fragments, can be separated on a Waters Protein-Pak Hi Res Q Column. The AEX separation result is highly consistent with the agarose gel separation. The size of a dsDNA can be assessed by comparing the retention time of the dsDNA with that of the 1 kb Plus DNA Ladder. This method can be used, but not limited, to separate and assess the size of the DNA fragments obtained from the restriction enzyme digestion of a plasmid. Such an analysis can serve as an assay to demonstrate the presence and the identity of the plasmid throughout the manufacturing process of gene therapy products.

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