Nick translation (or head translation), developed in 1977 by Peter Rigby and Paul Berg, is a tagging technique in

molecular biology Molecular biology is the branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions. The study of chemical and phys ...

in which

DNA Polymerase I DNA polymerase I (or Pol I) is an enzyme that participates in the process of prokaryotic DNA replication. Discovered by Arthur Kornberg in 1956, it was the first known DNA polymerase (and the first known of any kind of polymerase). It was initia ...

is used to replace some of the nucleotides of a DNA sequence with their labeled analogues, creating a tagged DNA sequence which can be used as a probe in

fluorescent in situ hybridization Fluorescence ''in situ'' hybridization (FISH) is a molecular cytogenetic technique that uses fluorescent probes that bind to only particular parts of a nucleic acid sequence with a high degree of sequence complementarity. It was developed by ...

(FISH) or blotting techniques. It can also be used for

radiolabeling A radioactive tracer, radiotracer, or radioactive label is a chemical compound in which one or more atoms have been replaced by a radionuclide so by virtue of its radioactive decay it can be used to explore the mechanism of chemical reactions by t ...

. This process is called nick translation because the DNA to be processed is treated with DNAase to produce single-stranded "nicks". This is followed by replacement in nicked sites by

DNA polymerase I DNA polymerase I (or Pol I) is an enzyme that participates in the process of prokaryotic DNA replication. Discovered by Arthur Kornberg in 1956, it was the first known DNA polymerase (and the first known of any kind of polymerase). It was initia ...

, which elongates the 3' hydroxyl terminus, removing nucleotides by 5'-3' exonuclease activity, replacing them with dNTPs. To radioactively label a DNA fragment for use as a probe in blotting procedures, one of the incorporated nucleotides provided in the reaction is radiolabeled in the ''alpha'' phosphate position. Similarly, a fluorophore can be attached instead for fluorescent labelling, or an antigen for immunodetection. When DNA polymerase I eventually detaches from the DNA, it leaves another nick in the phosphate backbone. The nick has "translated" some distance depending on the processivity of the polymerase. This nick could be sealed by

DNA ligase DNA ligase is a specific type of enzyme, a ligase, () that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. It plays a role in repairing single-strand breaks in duplex DNA in living orga ...

, or its 3' hydroxyl group could serve as the template for further DNA polymerase I activity. Proprietary enzyme mixes are available commercially to perform all steps in the procedure in a single incubation. Nick translation could cause double-stranded DNA breaks, if DNA polymerase I encounters another nick on the opposite strand, resulting in two shorter fragments. This does not influence the performance of the labelled probe in in-situ hybridization.

References

Biochemistry detection methods Genetics techniques Laboratory techniques Molecular biology techniques {{Genetics-stub