DNAase footprinting a simple method for the detection of protein-DNA binding specificity

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Abstract
A method for studying the sequence-specific binding of proteins to DNA is described. The technique is a simple conjoining of the Maxam-Gilbert DNA-sequencing method and the technique of DNAase-protected fragment isolation. Fragments of a 5′ end-labelled, double-stranded DNA segment, partially degraded by DNAase in the presence and absence of the binding protein, are visualized by eletrophoresis and autoradiography alongside the base-specific reaction products of the Maxam-Gilbert sequencing method. It is then possible to see the protective “footprint” of the binding protein on the DNA sequence. The binding of lac repressor to lac operator is visualized by “footprinting” as an example. Equilibrium estimates indicate that 10-fold sequence-specificity (differential binding constant) could be studied easily using this technique.