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Mobility Shift DNA‐Binding Assay Using Gel Electrophoresis

The DNA-binding assay using mobility shift polyacrylamide gel electrophoresis (pAGE) is a simple. rapid, and extremely sensitive method for the detection of sequence-specific DNA-binding proteins in crude extracts. This assay also permits the quantitative determination of the affinity, abundance, association rate constants, dissociation rate constants, and binding specificity of DNA-binding proteins. Proteins that bind specifically to an end-labeled DNA fragment retard the mobility of the fragment during electrophoresis, resulting in discrete bands corresponding to the individual protein-DNA complexes.