Nick translation

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As a nick translation is in molecular biology , a DNA - labeling technique called. The repair function of DNA polymerase I from Escherichia coli is used to incorporate marked nucleotides into single-strand breaks, so-called nicks .

DNA polymerase I is an enzyme that can repair single-strand breaks or single-strand gaps in the DNA, which can lead to transcription problems or even to the tearing of the DNA molecule due to mechanical stress . For this purpose, the polymerase has a 5 '→ 3' exonuclease activity in which the nucleotides are removed from the 5 'end in the direction of the 3' end. This exonuclease activity is a specialty of DNA polymerase I. It should not be confused with the 3 '→ 5' exonuclease activity for proofreading, which DNA polymerase I also has. The strand break is not closed (a DNA ligase is required for this), but shifted. Hence the name Nick Translation .

If nucleotides marked radioactively ( 3 H , α- 32 P ) or marked by a small molecule ( digoxygenin , biotin , fluorescent dye ) are added as a substrate , these are incorporated and mark a region of the DNA strand. The intensity of the marking depends on the number of strand breaks and this is determined by the concentration of added DNase I which causes the nicks . The detection of the marking depends on the marker. The detection takes place z. B. by a fluorescence microscope or blotting techniques.

This marking system was developed in 1977 by Peter Rigby and colleagues.

Individual evidence

  1. ↑ Rigby, PW et al. (1977): Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. In: J. Mol. Biol. Vol. 113, pp. 237-251. PMID 881736 doi : 10.1016 / 0022-2836 (77) 90052-3

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