Nick Translation and Random Hexamer Labeling of DNA

  • Jane Davenport-Jones
Part of the Springer Protocols Handbooks book series (SPH)


Labeled nucleotides (radioactive or fluorescent) can be incorporated efficiently into double-stranded DNA by nick translation. Nick translation works by using DNase and DNA polymerase I enzymes. DNase cuts one strand of the DNA, exposing 5′-phosphoryl and 3′-hydroxyl (OH) termini DNA. Polymerase I adds deoxyribonucleoside triphosphate (dNTPs), including labeled dNTPs to the exposed 3′-OH strand and at the same time, the polymerase exonuclease activity digests from the exposed 5′ end. In this way, a new complementary strand, including labeled dNTPs is produced (1). It is also possible to incorporate radioactive nucleotides into DNA using a enzymatic primer extension technique (2). In this method, random hexanucleotides are annealed to the denatured DNA to be used as the probe. These are used as a primer for enzymatic extension in the presence of the four dNTPs, one of which is radiolabeled.


Amersham Pharmacia Biotech Stop Solution Complementary Strand Nick Translation Unincorporated Nucleotide 
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  1. 1.
    Rigby, P. W. J., Dieckmann, M., Rhodes, C., and Berg, P. (1977) Labelling deoxyribo-nucleic acid to a high specific activity in vitro by nick translation with DNA polymerase I. J. Mol. Biol. 113, 237–251.PubMedCrossRefGoogle Scholar
  2. 2.
    Feinberg, A. P. and Vogelstein, B. (1983) A technique for radiolabelling DNA restriction endonuclease fragments to a high specific activity. Anal. Biochem. 132, 6–13.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2000

Authors and Affiliations

  • Jane Davenport-Jones
    • 1
  1. 1.Department of BiosciencesUniversity of HertfordshireHafieldUK

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