DNase Footprinting Assay

DNase footprinting assay ( DNase footprint assay ) is a molecular technique for the detection of DNA - protein interactions. The fact is utilized that the DNA is protected at locations where a protein is bound to a certain extent before enzymatic cleavages. In the method, the enzyme is deoxyribonuclease (abbreviated DNase ) was used. It ensures that DNA chains are cleaved by the enzyme once or a few places.

The investigated DNA chains radiolabeled at the end of the two strands. They are added to a sample of cell protein extract or a sample of specifically selected proteins. It may form DNA-protein bindings. The addition of DNase arise DNA fragments of different length. In particular, those fragments of any length are interesting, containing the labeled end, because only they are visible in the subsequent gel electrophoresis. After effect of DNase proteins of the DNA are separated. In the gel electrophoresis results in a nearly continuous distribution of the fragments, as the speed of a DNA fragment is proportional to its length and should occur any length. Only in sections on which proteins were bound, DNase could not act. DNA fragments whose length falls within this binding region is missing, which is visible through a gap of the continuous propagation in the gel. Such a gap is called the footprint of the protein to the DNA. The sequence of the DNA binding protein can be reconstructed from the marked end of the basis of the known distance between the binding site.