Document Detail


Electrophoretic mobility shift assay (EMSA) for detecting protein-nucleic acid interactions.
MedLine Citation:
PMID:  17703195     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
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.
Authors:
Lance M Hellman; Michael G Fried
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural    
Journal Detail:
Title:  Nature protocols     Volume:  2     ISSN:  1750-2799     ISO Abbreviation:  Nat Protoc     Publication Date:  2007  
Date Detail:
Created Date:  2007-08-17     Completed Date:  2007-12-06     Revised Date:  2014-09-16    
Medline Journal Info:
Nlm Unique ID:  101284307     Medline TA:  Nat Protoc     Country:  England    
Other Details:
Languages:  eng     Pagination:  1849-61     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Bacterial Proteins / metabolism
Cyclic AMP Receptor Protein / metabolism
DNA-Binding Proteins / analysis*,  metabolism
Electrophoretic Mobility Shift Assay / methods*
Escherichia coli / genetics,  metabolism
Escherichia coli Proteins / metabolism
Humans
Lac Operon / genetics
Lac Repressors
O(6)-Methylguanine-DNA Methyltransferase / metabolism
Promoter Regions, Genetic
RNA-Binding Proteins / analysis*
Repressor Proteins / metabolism
Transcription Factors / metabolism
Grant Support
ID/Acronym/Agency:
GM-070662/GM/NIGMS NIH HHS; R01 GM070662/GM/NIGMS NIH HHS; R01 GM070662-01/GM/NIGMS NIH HHS; R01 GM070662-02/GM/NIGMS NIH HHS; R01 GM070662-03/GM/NIGMS NIH HHS; R01 GM070662-04/GM/NIGMS NIH HHS; R01 GM070662-05/GM/NIGMS NIH HHS; R01 GM070662-06/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Bacterial Proteins; 0/Cyclic AMP Receptor Protein; 0/DNA-Binding Proteins; 0/Escherichia coli Proteins; 0/Lac Repressors; 0/LacI protein, E coli; 0/RNA-Binding Proteins; 0/Repressor Proteins; 0/Transcription Factors; 0/crp protein, E coli; EC 2.1.1.63/O(6)-Methylguanine-DNA Methyltransferase
Comments/Corrections

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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