Antonie van Leeuwenhoek

, Volume 36, Issue 1, pp 461–474 | Cite as

Evaluation of membrane filter methods for DNA-DNA hybridization

  • J. De Ley
  • R. Tijtgat


Three membrane filter methods for DNA hybridization were compared. In all of them filter-fixed DNA is spontaneously released during incubation. The effect depends largely on the temperature: 40 to 85% of DNA is released at 70 to 80 C; at higher temperatures most of the DNA is released and heterologous DNA relatedness values are unreliable. Only the dimethylsulfoxide filter method (Legault-Démare et al., 1967) can be recommended for the entire temperature and % GC range.

The methods of Denhardt (1966) and of Warnaar and Cohen (1966) are not recommended for hybridizations of DNA containing more than 50% GC. Impure DNA yields too low DNA relatedness values.


Dimethylsulfoxide Membrane Filter Filter Method Membrane Filter Method 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Bonner, J., Kung, G. andBekhor, I. 1967. A method for the hybridization of nucleic acid molecules at low temperatures. - Biochemistry6: 3650–3653.Google Scholar
  2. Bray, G. A. 1960. A simple efficient liquid scintillator for counting aqueous solutions in a liquid scintillator counter. - Anal. Biochem.1: 279–285.Google Scholar
  3. Brenner, D. J., Fanning, G. R., Johnson, K. E., Citarella, R. V. andFalkow, S. 1969. Polynucleotide sequence relationships among members of Enterobacteriaceae. - J. Bacteriol.98: 637–650.Google Scholar
  4. De Ley, J., Cattoir, H. andReynaerts, A. 1970. The quantitative measurement of DNA hybridization from renaturation rates. - Europ. J. Biochem.12: 133–142.Google Scholar
  5. De Ley, J., Park, I. W., Tijtgat, R. andVan Ermengem, J. 1966. DNA homology and taxonomy ofPseudomonas andXamhomonas. - J. Gen. Microbiol.42: 43–56.Google Scholar
  6. De Ley, J. andVan Muylem, J. 1963. Some applications of deoxyribonucleic acid base composition in bacterial taxonomy. - Antonie van Leeuwenhoek29: 344–358.Google Scholar
  7. Denhardt, D. T. 1966. A membrane-filter technique for the detection of complementary DNA. - Biochem. Biophys. Res. Comm.23: 641–646.Google Scholar
  8. Gillespie, D. 1968. The formation and detection of DNA-RNA hybrids, p. 641–668.In S. P. Colowick and N. O. Kaplan, (eds.), Methods in Enzymology, Vol. XII B. - Acad. Press, New York.Google Scholar
  9. Gillespie, D. andSpiegelman, S. 1965. A quantitative assay for DNA-RNA hybrids with DNA immobilized on a membrane. - J. Mol. Biol.12: 829–842.Google Scholar
  10. Gillis, M., De Ley, J. andDe Cleene, M. 1970. The determination of molecular weight of bacterial genome DNA from renaturation rates. - Europ. J. Biochem.12: 143–153.Google Scholar
  11. Heberlein, G. T., De Ley, J. andTijtgat, R. 1967. Deoxyribonucleic acid homology and taxonomy ofAgrobacterium, Rhizobium andChromobacterium. - J. Bacteriol.94: 116–124.Google Scholar
  12. Hoyer, B. H., McCarthy, B. J. andBolton, E. T. 1964. A molecular approach in the systematics of higher organisms. - Science144: 959–967.Google Scholar
  13. Johnson, J. L. andOrdal, E. J. 1968. Deoxyribonucleic acid homology in bacterial taxonomy: effect of incubation temperature on reaction specificity. - J. Bacteriol.95: 893–900.Google Scholar
  14. Kingsbury, D. T., Fanning, G. R., Johnson, K. E. andBrenner, D. J. 1969. Thermal stability of interspeciesNeisseria DNA duplexes. - J. Gen. Microbiol.55: 201–208.Google Scholar
  15. Kirby, K. S. 1957. A new method for the isolation of deoxyribonucleic acids: evidence on the nature of bonds between deoxyribonucleic acid and protein. - Biochem. J.66: 495–504.Google Scholar
  16. Kirby, K. S., Fox-Carter, E. andGuest, M. 1967. Isolation of deoxyribonucleic acid and ribosomal ribonucleic acid from bacteria. - Biochem. J.104: 258–262.Google Scholar
  17. Legault-Démare, J., Desseaux, B., Heyman, T., Séror, S. andRess, G. P. 1967. Studies on hybrid molecules of nucleic acids. 1. DNA-DNA hybrids on nitrocellulose filters. - Biochem. Biophys. Res. Comm.28: 550–557.Google Scholar
  18. Marmur, J. 1961. A procedure for the isolation of deoxyribonucleic acid from micro-organisms. - J. Mol. Biol.3: 208–218.Google Scholar
  19. Marmur, J. andDoty, P. 1961. Thermal renaturation of deoxyribonucleic acids. - J. Mol. Biol.3: 585–594.Google Scholar
  20. McCarthy, B. J. 1967. Arrangement of base sequences in deoxyribonucleic acid. - Bacteriol. Rev.31: 215–229.Google Scholar
  21. Van den Berghe, C., Schnek, G. etLedoux, L. 1969. Remarques sur les techniques d'hybridation DNA-DNA. - Arch. int. Physiol. Biochem.77: 176.Google Scholar
  22. Warnaar, S. O. andCohen, J. A. 1966. A quantitative assay for DNA-DNA hybrids using membrane filters. - Biochem. Biophys. Res. Comm.24: 554–558.Google Scholar

Copyright information

© Swets & Zeitlinger 1970

Authors and Affiliations

  • J. De Ley
    • 1
  • R. Tijtgat
    • 1
  1. 1.Laboratory for Microbiology and Microbial Genetics, Faculty of SciencesState UniversityGentBelgium

Personalised recommendations