Detection of RNA-Protein Complexes by Electrophoretic Mobility Shift Assay

The electrophoretic mobility shift assay (EMSA) is commonly used to study protein-nucleic acid interactions. The technique is based on the observation that protein-bound nucleic acid molecules migrate more slowly than free nucleic acid molecules when subjected to native polyacrylamide or agarose gel electrophoresis. After electrophoresis, the distribution of species containing nucleic acid is determined by autoradiography or other sensitive imaging technique. Under appropriate conditions, the method can be used for quantitative binding analysis, but it is most widely used as a qualitative means of detecting protein-RNA complexes. In this chapter, features relevant to the design of EMSA experiments are identified, and the technical factors found to be important for the success of the assay are outlined.