Size and Purity Assessment of Single-Guide RNAs by Anion-Exchange Chromatography (AEX)

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Hua Yang Stephan M. Koza Ying Qing Yu
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Application Notes
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This application note demonstrates anion-exchange chromatography is robust, reproducible, easy to automate, yields quantitative information, and requires a small amount of sample. We demonstrate here that the components of a Low Range ssRNA Ladder, ranging from 50 to 500 bases, can be separated on a Waters Protein-Pak Hi Res Q Column with a linear correlation between the log of base-number and observed retention time when TMAC is used as an elution salt. The size of ssRNAs ranging from 100 to 150 bases can be estimated by comparing the retention time of the ssRNAs with that of the Low Range ssRNA Ladder. In addition, the purity of a sgRNAs may also be observed from the same chromatographic separation. This method can potentially be applied to the analysis of sgRNAs which are the key element for CRISPR/Cas9 gene editing technology.

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