Article

Plant Molecular Biology Reporter

, Volume 16, Issue 1, pp 49-59

First online:

DNA Isolation and AFLP™ Genetic Fingerprinting of shape Theobroma cacao (L.)

  • Matthew D. PerryAffiliated withPlant Science Division, School of Biological Sciences, University of Nottingham
  • , Michael R. DaveyAffiliated withPlant Science Division, School of Biological Sciences, University of Nottingham
  • , J. Brian PowerAffiliated withPlant Science Division, School of Biological Sciences, University of Nottingham
  • , Kenneth C. LoweAffiliated withPlant Science Division, School of Biological Sciences, University of Nottingham
  • , H. Frances J. BlighAffiliated withPlant Science Division, School of Biological Sciences, University of Nottingham
  • , Paul S. RoachAffiliated withSchool of Biomedical Sciences, Queens Medical Centre, University of Nottingham
  • , Chris JonesAffiliated withSchool of Biomedical Sciences, Queens Medical Centre, University of Nottingham

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Abstract

Accurate identification of parental plants and their hybrids is essential for an effective breeding programme. Traditional classification of cocoa varieties relies on the characterisation of agricultural traits at plant maturity. A rapid and reliable method is described, based on genotypic analysis. An efficient DNA isolation procedure was developed, yielding unsheared DNA of high purity. Two genetic fingerprinting techniques, RAPD and AFLP™, were evaluated for their suitability in distinguishing cocoa varieties. RAPD analysis was unsatisfactory due to the low frequency of polymorphisms and poor reproducibility. AFLP™ was reliable in distinguishing phenotypically identical, known varieties of cocoa. Importantly, AFLP™ also revealed intra- and inter-varietal variation.

Abbreviations: AFLP™, amplified fragment length polymorphism; APS, ammonium persulphate; CTAB, hexadecyltrimethylammonium bromide; DEB, DNA extraction buffer; f.wt., fresh weight; NEB, nuclei extraction buffer; PMSF, phenylmethanesulphonyl fluoride; RAPD, random amplified polymorphic DNA; T4 PNK, Bacteriophage T4 polynucleotide kinase; Taq, Thermus aquaticus; TBE, tris-borate-EDTA; TEMED, NNN′N′ tetramethylethylenediamine.

amplified fragment length polymorphism cocoa RAPD woody plant