An electrophoretic mobility shift assay (EMSA) or mobility shift electrophoresis, also referred as a gel shift assay, gel mobility shift assay, band shift assay, or gel retardation assay, is a common affinity electrophoresis technique used to study protein–DNA or protein–RNA interactions. This procedure can determine if a protein . This article provides information about studying DNA–protein interactions using an electrophoretic mobility shift assay (EMSA). Introduction to the EMSA.
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 . EMSA also can be used to study protein- or sequence-dependent DNA bending. The electrophoretic mobility shift assay (EMSA) was developed by Garner and Revzin and Fried and Crothers. Excellent treatments of the theory underlying this assay system may be found elsewhere. We note that we have yet to find conditions under which activated Spo0A shifts the mobility of DNA fragments . Electrophoretic mobility shift assay is a very important technique to understand DNA – protein interaction.
If you have a fluorescence reader device, the best is simply to fluorescently-label your probe (eg with TAMRA or Cy3) and perform the detection directly on gel. No need to use expensive kits, or radioactivity.
When a large molar excess of unlabeled competitor DNA is adde the mobility shift is greatly reduced. The EMSA ( electrophoretic mobility shift assay ) is used to study. IRDye 7oligos are end-labeled on both strands. This is a sample Mobility Shift Protocol (NFκB). See the specific EMSA oligo pack insert for more informati.
Currently I am performed EMSA assays coupled with SC and NSC competitors. I place my binding buffer, recombinant protein, and other . I have recombinant protein that I am using. EMSA originally used widely in the study of sequence-specific DNA-binding proteins such as transcription factors, has been further developed to investigate DNA-protein interactions . The Gelshift Chemiluminescent EMSA Assay Kit provides a simple, non- radioactive assay to identify protein-DNA binding with proven reagents.
In this electrophoretic mobility shift assay (EMSA), cell extracts or purified factors are incubated with biotin end-labeled probe containing the consensus binding site of interest. It is originally used to detect transcription factors, and is now further developed into investigating DNA-protein interactions, RNA-protein interactions, . This assay can be used to determine, in both a qualitative and quantitative manner, if a particular transcription factor is present within . More than 4EMSA kits now available covering most widely studied transcription factors. Affymetrix EMSA ( Electrophoretic Mobility Shift Assay ) Kits are useful tools for .