The Histochemical Journal

, Volume 22, Issue 12, pp 639–652

Patterns of digestion of human chromosomes by restriction endonucleases demonstrated byin situ nick translation

  • A. T. Sumner
  • M. H. Taggart
  • R. Mezzanotte
  • L. Ferrucci
Papers

DOI: 10.1007/BF01047448

Cite this article as:
Sumner, A.T., Taggart, M.H., Mezzanotte, R. et al. Histochem J (1990) 22: 639. doi:10.1007/BF01047448

Summary

A restriction enzyme-nick translation procedure has been developed for localizing sites of restriction endonuclease action on chromosomes. This method involves digestion of fixed chromosome preparations with a restriction enzyme, nick translation with DNA polymerase I in the presence of biotinylated-dUTP, detection of the incorporated biotin label with streptavidinalkaline phosphatase, and finally staining for alkaline phosphatase. Results obtained on human chromosomes using a wide variety of restriction enzymes are described, and compared with results of Giemsa and Feulgen staining after restriction enzyme digestion. Results of nick translation are not in general the opposite of those obtained with Giemsa staining, as might have been expected. Although the nick translation procedure is believed to give a more accurate picture of the distribution of restriction enzyme recognition sites on chromosomes than Giemsa staining, it is clear that the results of the nick translation experiments are affected by accessibility to the enzymes of the chromosomal DNA, as well as by the extractability of the DNA.

Copyright information

© Chapman and Hall Ltd 1990

Authors and Affiliations

  • A. T. Sumner
    • 1
  • M. H. Taggart
    • 1
  • R. Mezzanotte
    • 2
  • L. Ferrucci
    • 3
  1. 1.MRC Human Genetics UnitWestern General HospitalEdinburghScotland, UK
  2. 2.Istituto di Biologia Generale, Facolta di Medicina e ChirurgiaUniversita di CagliariCagliariItaly
  3. 3.Dipartimento di BiologiaII Universita degli Studi di RomaRomaItaly