Two-dimensional gel selection of protein binding sites on DNA.
We have developed a gel electrophoretic approach for visualizing and cloning protein binding sites from complete genomes. This system consists of a simple two-dimensional band shift, in which protein-DNA complexes are retarded in the first dimension, performed at low temperature, and disrupted in the second dimension, performed at high temperature. We present here results obtained with the integration host factor (IHF) and cAMP receptor protein (CRP) proteins of Escherichia coli, and discuss some of the important methodological aspects of the technique.
Boffini
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A
Department of Molecular Biology, University of Geneva, Switzerland.
Prentki
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P
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D002373:Cyclic AMP Receptor Protein / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000737:chemistry; D034982:Integration Host Factors; D008969:Molecular Sequence Data
D001426:Bacterial Proteins; D002373:Cyclic AMP Receptor Protein; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D034982:Integration Host Factors