Hox Genes pp 211-230 | Cite as

Measuring Hox-DNA Binding by Electrophoretic Mobility Shift Analysis

  • Kelly Churion
  • Ying Liu
  • Hao-Ching Hsiao
  • Kathleen S. Matthews
  • Sarah E. BondosEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1196)


Understanding gene regulation by Hox transcription factors requires understanding the forces that underlie DNA binding by these proteins. Electrophoretic mobility shift analysis (EMSA) not only allows measurement of protein affinity and cooperativity but also permits visualization of differently migrating protein-DNA complexes, including complexes with different compositions or complexes with identical compositions yet assembled in different geometries. Furthermore, protein activity can be measured, allowing correction of binding constants for the percentage of protein that is properly folded and capable of binding DNA. Protocols for measuring protein activity and the equilibrium DNA-binding dissociation constant (K d) are provided. This versatile assay system can be adjusted based on specific needs to measure other parameters, including the kinetic association and dissociation constants (k a and k d) and the formation of heterologous protein-protein interactions.

Key words

Hox Gel retardation Gel shift Electrophoretic mobility shift analysis EMSA DNA binding Activity Affinity Cooperativity 



This work was supported by an RDEAP grant from the Texas A&M Health Science Center to S.E.B. and a Robert A. Welch Foundation grant (C-576) to K.S.M.


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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Kelly Churion
    • 1
  • Ying Liu
    • 2
  • Hao-Ching Hsiao
    • 1
  • Kathleen S. Matthews
    • 3
  • Sarah E. Bondos
    • 4
    • 3
    Email author
  1. 1.Department of Molecular and Cellular MedicineTexas A&M Health Science CenterCollege StationUSA
  2. 2.Medical Genomics Laboratory, UAB School of MedicineUniversity of Alabama at BirminghamBirminghamUSA
  3. 3.Department of Biochemistry and Cell BiologyRice UniversityHoustonUSA
  4. 4.Department of Molecular and Cellular MedicineTexas A&M Health Science CenterCollege StationUSA

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