Labeling of DNA Probes by Nick Translation

In this method, E. coli DNA Pol I binds to a nick or short gap in duplex DNA. The 5′ → 3′ exonuclease activity of Pol I then removes nucleotides from one strand of the DNA, creating a template for the synthesis of DNA by the 5′ → 3′ polymerase activity of Pol I. The simultaneous elimination of nucleotides from the 5′ side and the addition of nucleotides to the 3′ side result in movement of the nick (nick translation) along the DNA, which becomes labeled to high specific activity.