Electrophoretic mobility shift assay (EMSA) for detecting protein-nucleic acid interactions

Lance M. Hellman, Michael G. Fried

Research output: Contribution to journalArticlepeer-review

754 Scopus citations

Abstract

The gel electrophoresis mobility shift assay (EMSA) is used to detect protein complexes with nucleic acids. It is the core technology underlying a wide range of qualitative and quantitative analyses for the characterization of interacting systems. In the classical assay, solutions of protein and nucleic acid are combined and the resulting mixtures are subjected to electrophoresis under native conditions through polyacrylamide or agarose gel. After electrophoresis, the distribution of species containing nucleic acid is determined, usually by autoradiography of 32P-labeled nucleic acid. In general, protein-nucleic acid complexes migrate more slowly than the corresponding free nucleic acid. In this protocol, we identify the most important factors that determine the stabilities and electrophoretic mobilities of complexes under assay conditions. A representative protocol is provided and commonly used variants are discussed. Expected outcomes are briefly described. References to extensions of the method and a troubleshooting guide are provided.

Original languageEnglish
Pages (from-to)1849-1861
Number of pages13
JournalNature Protocols
Volume2
Issue number8
DOIs
StatePublished - Aug 2007

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology (all)

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