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Nucleic Acids Research, 2000, Vol. 28, No. 24 e105
© 2000 Oxford University Press

TE-AFLP: combining rapidity and robustness in DNA fingerprinting

A. W. G. van der Wurff*, Y. L. Chan1, N. M. van Straalen and J. Schouten1

Institute of Ecological Science, Faculty of Biology, De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands and 1MRC Holland, Hudsonstraat 68, 1057 SN Amsterdam, The Netherlands

A new type of fingerprinting technique is presented, based on amplified fragment length polymorphism (AFLP). Rather than two endonucleases as in AFLP, we propose the use of three enzymes, hence the method is called three endonuclease (TE)-AFLP. Genomic DNA is digested and two sets of adapters are selectively ligated onto the restriction fragments in a single reaction volume. No adapters complementary to the ends generated by a frequent cutter are added. Due to the addition of a third endonuclease, the TE-AFLP method provides a high discriminatory power and a reduction in the number of bands. The latter makes it especially suitable for the analysis of complex genomes. TE-AFLP fingerprints are suitable for detection by automatic fluorescent sequencers and are obtained in less than half the time and at reduced costs compared to a typical AFLP. The reliability of this method was investigated by determining the influence of varying digestion, ligation and PCR components on the fingerprint. Moreover, cross-experiments to study inheritance of loci were performed with a primitive insect and with tomato strains. The features of TE-AFLP are discussed in comparison with conventional AFLP.

* To whom correspondence should be addressed. Tel: +31 20 444 7073; Fax: +31 20 444 7123; Email: wurff@bio.vu.nl


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