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AFLP technology for DNA fingerprinting

Nature Protocols volume 2, pages 1387–1398 (2007)Cite this article

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Abstract

The AFLP technique is a powerful DNA fingerprinting technology applicable to any organism without the need for prior sequence knowledge. The protocol involves the selective PCR amplification of restriction fragments of a total digest of genomic DNA, typically obtained with a mix of two restriction enzymes. Two limited sets of AFLP primers are sufficient to generate a large number of different primer combinations (PCs), each of which will yield unique fingerprints. Visualization of AFLP fingerprints after gel electrophoresis of AFLP products is described using either a conventional autoradiography platform or an automated LI-COR system. The AFLP technology has been used predominantly for assessing the degree of variability among plant cultivars, establishing linkage groups in crosses and saturating genomic regions with markers for gene landing efforts. AFLP fragments may also be used as physical markers to determine the overlap and positions of genomic clones and to integrate genetic and physical maps. Crucial characteristics of the AFLP technology are its robustness, reliability and quantitative nature. This latter feature has been exploited for co-dominant scoring of AFLP markers in sample collections such as F2 or back-cross populations using appropriate AFLP scoring software. This protocol can be completed in 2–3 d.

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Figure 1: Outline of the AFLP procedure.
Figure 2: Schematic for adapter and primer design for the two rare cutters EcoRI and PstI and the frequent cutter MseI.
Figure 3: AFLP analysis of 32 tomato F2 segregants.
Figure 4: Gel image of an AFLP analysis of 56 Arabidopsis Recombinant Inbred Line (RIL) segregants analyzed using an EcoRI + AA/MseI + CAA primer combination.

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Change history

  • 14 August 2008

    In the version of this article initially published, the three genotype classes in Figure 3b were incorrectly described in the legend. "A: homozygous as the first parent; H: heterozygous, B: homozygous as the second parent" should have read "A: homozygous absent; H: heterozygous; B:homozygous present". This error has been corrected in the HTML and PDF versions of the article.

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Acknowledgements

We are grateful to H. Van den Daele and I. Vercauteren for their help with the manuscript. The AFLP technology is covered by patents and patent applications owned by Keygene N.V. AFLP and AFLP-QuantarPro are registered trademarks of Keygene N.V. All other product names, brand names or company names are used for identification purposes only and may be (registered) trademarks of their respective owners.

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Authors and Affiliations

  1. Department of Plant Systems Biology, VIB, Technologiepark 927, Ghent, B-9052, Belgium

    Marnik Vuylsteke

  2. Department of Molecular Genetics, Ghent University, Technologiepark 927, Ghent, B-9052, Belgium

    Marnik Vuylsteke

  3. Keygene N.V., Agro Business Park 90, Wageningen, NL-6708, PW, The Netherlands

    Johan D Peleman & Michiel JT van Eijk

Authors
  1. Marnik Vuylsteke
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  3. Michiel JT van Eijk
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Correspondence to Marnik Vuylsteke.

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Competing interests

M.J.T. van Ejik and J.D. Peleman are full-time employees of Keygene N.V.

M. Vuylsteke declares not to have competing interests.

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Vuylsteke, M., Peleman, J. & van Eijk, M. AFLP technology for DNA fingerprinting. Nat Protoc 2, 1387–1398 (2007). https://doi.org/10.1038/nprot.2007.175

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