Plant Molecular Biology Reporter

, Volume 24, Issue 1, pp 81–91 | Cite as

A rapid method for tissue collection and high-throughput isolation of genomic DNA from mature trees

  • Josquin F. G. Tibbits
  • Luke J. McManus
  • Antanas V. Spokevicius
  • Gerd BossingerEmail author


Collection of tissue and subsequent isolation of genomic DNA from mature tree species often proves difficult. DNA extraction from needles, leaves, or buds is recommended in many protocols. Collecting these tissues from mature trees generally requires the use of firearms or climbing if sampling is to be nondestructive. As a result, sample collection is a major expense of many tree-based projects. Tree (and plant) tissues generally contain large amounts of polysaccharides and phenolic compounds that are difficult to separate from DNA. Many methods aim to overcom these problems, with most involving extraction in buffers containing the nonionic detergent cetyltrimethyl-ammonium bromide (CTAB), followed by numerous steps to clean contaminants from the DNA, using organic solvents and differential salt precipitation. These steps are time-consuming, such that isolation of DNA becomes the bottleneck in many molecular studies. This paper presents a new, efficient, cambium collection method for tree species and a DNA extraction protocol based on that of Doyle and Doyle (1987), with follow-up purification using the Wizard nuclei lysis and protein precipitation solutions (Promega). Results show a significant improvement in yield and DNA purity compared with other published methods, with consistently high yields of pure genomic DNA and high sample throughput. The relatively low cost per extraction, no requirement for use of liquid nitrogen, no requirement for freezer storage, and long-term sample stability after collection are important additional benefits.

Key words

DNA extraction cambial scrapings Eucalyptus Pinus tree sample collection 



bovine serum albumin


cetyltrimethylammonium bromide


poly vinylpyrrollidone


tris-EDTA buffer


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  1. Bousquet J, Simpson L, and Lalonde M (1990) DNA amplification from vegetative and sexual tissues of trees using polymerase chain reaction. Can J For Res 20: 254–257.CrossRefGoogle Scholar
  2. Carlson JE, Tulsieram LK, Glaubitz JC, Luk VWK, Kauffeldt C, and Rutleidge R (1991) Segregation of random amplified DNA markers in F1 progeny of conifers. Theor Appl Genet 83: 194–200.CrossRefGoogle Scholar
  3. Chang S, Puryear J, and Cairney J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Mol Biol Rep 11: 113–116.CrossRefGoogle Scholar
  4. Dellaporta SL, Wood J, and Hicks JB (1983) A plant DNA minipreparation version II. Plant Mol Biol Rep 1: 19–21.CrossRefGoogle Scholar
  5. Devey M, Jermstad KD, Tauer CG, and Neale DB (1991) Inheritance of RFLP loci in a loblolly pine three-generation pedigree. Theor Appl Genet 83: 238–242.CrossRefGoogle Scholar
  6. Devey M, Bell JC, Smith DN, Neale DB, and Moran GF (1996) A genetic linkage map forPinus radiata based on RFLP, RAPD, and microsatellite markers. Theor Appl Genet 92: 673–679.CrossRefGoogle Scholar
  7. Doyle JJ and Doyle JH (1987) Isolation of plant DNA from fresh tissue. Focus 12: 13–15.Google Scholar
  8. Jobes DV, Hurley DL, and Thien LB (1995) Plant DNA isolation: method to efficiently remove polyphenolics, polysaccharides and RNA. Taxon 44: 379–386.CrossRefGoogle Scholar
  9. Katterman FRH and Shattuck VI (1983) A effective method of DNA isolation from the mature leaves ofGossypium species that contain large amounts of phenolic tarpenoids and tannins. Prep Biochem 13: 347–359.PubMedCrossRefGoogle Scholar
  10. Kim CS, Lee CH, Shin JS, Chung YS, and Hyung NI (1997) A simple and rapid method for isolation of high quality genomic DNA from fruit trees and conifers using PVP. Nucleic Acids Res 25: 1085–1086.PubMedCrossRefGoogle Scholar
  11. Kreader CA (1996) Relief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein. Appl Environ Microbiol 62: 1102–1106.PubMedGoogle Scholar
  12. Lin J-J and Kuo J (1998) A new reagent for simple isolation of plant genomic DNA. Focus 20: 46–48.Google Scholar
  13. Mannerlöf M and Tenning P (1997) Screening of transgenic plants by multiplex PCR. Plant Mol Biol Rep 15: 38–45.CrossRefGoogle Scholar
  14. Murray CG and Thompson WF (1980) Rapid isolation of high molecular weight plant DNA. Nucleic Acids Res 8: 4321–4325.PubMedCrossRefGoogle Scholar
  15. Nelson CD, Kubisiak TL, and Stine M (1994) A genetic linkage map of longleaf pine (Pinus palustris Mill.) based on random amplified polymorphic DNA. J Hered 85: 433–439.Google Scholar
  16. Ostrowska E, Muralitharan M, Chandler S, Volker P, Hetherington S, and Dunshea F (1998) Technical review: optimizing conditions for DNA isolation fromPinus radiata. In Vitro Cell Dev Biol Plant 34: 108–111.CrossRefGoogle Scholar
  17. Rogers SO and Bendich AJ (1985) Extraction of DNA from milligram amounts of fresh, herbarium and mummified plant tissues. Plant Mol Biol 5: 69–76.CrossRefGoogle Scholar
  18. Sambrook J, Fritsch EF, and Maniatis T (1989) Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, New York.Google Scholar
  19. Stewart NC and Via LE (1993) A rapid CTAB DNA isolation technique useful for RAPD fingerprinting and other PCR applications. Biotechniques 14: 748–749.PubMedGoogle Scholar
  20. Wagner DB, Furnier GR, Saghai-Maroof MA, Williams SM, Dancik BP, and Allard RW (1987) Chloroplast DNA polymorphisms in lodgepole and jack pines and their hybrids. Proc Natl Acad Sci USA 84: 2097–2100.PubMedCrossRefGoogle Scholar

Copyright information

© Springer 2006

Authors and Affiliations

  • Josquin F. G. Tibbits
    • 1
  • Luke J. McManus
    • 1
  • Antanas V. Spokevicius
    • 1
  • Gerd Bossinger
    • 1
    Email author
  1. 1.School of Forest and Ecosystem ScienceThe University of MelbourneCreswickAustralia

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