Advertisement

Nuclear Ribosomal RNA Genes: ITS Region

  • Pascale Besse
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1115)

Abstract

Despite possible drawbacks (intraspecific polymorphisms and possible fungal contamination), sequencing of the ribosomal RNA gene ITS region remains one of the most popular nuclear sequences used for plant taxonomy and phylogeny. A protocol for PCR amplification and sequencing of this region using universal plant primers is provided.

Key words

Ribosomal DNA ITS Sequencing PCR 

Notes

Acknowledgements

Denis Da Silva and Michel Grisoni (UMR PVBMT Cirad-Université de la Réunion) are acknowledged for their help in the ITS amplification protocol design.

References

  1. 1.
    Baldwin BG, Sanderson MJ, Porter JM, Wojciechowski MF, Campbell CS, Donoghue MJ (1995) The ITS region of nuclear ribosomal DNA: a valuable source of evidence on angiosperm phylogeny. Ann Mo Bot Gard 82:247–277CrossRefGoogle Scholar
  2. 2.
    Hillis DM, Dixon MT (1991) Ribosomal DNA: molecular evolution and phylogenetic inference. Q Rev Biol 66:411–453PubMedCrossRefGoogle Scholar
  3. 3.
    Poczai P, Hyvönen J (2010) Nuclear ribosomal spacer regions in plant phylogenetics: problems and prospects. Mol Biol Rep 37:1897–1912PubMedCrossRefGoogle Scholar
  4. 4.
    Arnheim N (1983) Concerted evolution in multigene families. In: Nei M, Koehn R (eds) Evolution of genes and proteins. Sinauer, Sunderland, MA, pp 38–61Google Scholar
  5. 5.
    Dover G (1994) Concerted evolution, molecular drive and natural selection. Curr Biol 4:1165–1166PubMedCrossRefGoogle Scholar
  6. 6.
    Rogers SO, Bendich AJ (1987) Ribosomal RNA genes in plants: variability in copy number and in the intergenic spacer. Plant Mol Biol 9:509–520PubMedCrossRefGoogle Scholar
  7. 7.
    Alvarez IA, Wendel JF (2003) Ribosomal ITS sequences and plant phylogenetic inference. Mol Phylogenet Evol 29:417–434PubMedCrossRefGoogle Scholar
  8. 8.
    Buckler ES, Ippolito A, Holtsford TP (1997) The evolution of ribosomal DNA divergent paralogues and phylogenetic implications. Genetics 145:821–832PubMedGoogle Scholar
  9. 9.
    Calonje M, Martın-Bravo S, Dobes C, Gong W, Jordon-Thaden I, Kiefer C, Kiefer M, Paule J, Schmickl R, Koch MA (2009) Non-coding nuclear DNA markers in phylogenetic reconstruction. Plant Syst Evol 282:257–280CrossRefGoogle Scholar
  10. 10.
    Bachmann K (1994) Molecular markers in plant ecology Tansley review no. 63. New Phytol 126:403–418CrossRefGoogle Scholar
  11. 11.
    Zimmer EA, Wen J (2013) Using nuclear gene data for plant phylogenetics: progress and prospects. Mol Phylogenet Evol 66:539–550PubMedCrossRefGoogle Scholar
  12. 12.
    Feliner GN, Rosselló JA (2007) Better the devil you know? Guidelines for insightful utilization of nrDNA ITS in species-level evolutionary studies in plants. Mol Phylogenet Evol 44:911–919CrossRefGoogle Scholar
  13. 13.
    Besnard G, Rubio de Casas R, Vargas P (2007) Plastid and nuclear DNA polymorphism reveals historical processes of isolation and reticulation in the olive tree complex (Olea europaea). J Biogeogr 34:736–752CrossRefGoogle Scholar
  14. 14.
    Bailey CD, Carr TG, Harris SA, Hughesa CE (2003) Characterization of angiosperm nrDNA polymorphism, paralogy, and pseudogenes. Mol Phylogenet Evol 29:435–455PubMedCrossRefGoogle Scholar
  15. 15.
    Kress WJ, Wurdack KJ, Zimmer EA, Weig LA, Janzen DH (2005) Use of DNA barcodes to identify flowering plants. PNAS 102:8369–8374PubMedCrossRefGoogle Scholar
  16. 16.
    CBOL Plant Working Group (2009) A DNA barcode for land plants. PNAS 106:12794–12797CrossRefGoogle Scholar
  17. 17.
    Yao H, Song J, Liu C, Luo K, Han J, Li Y, Pang X, Xu H, Zhu Y, Xiao P, Chen S (2010) Use of ITS2 region as the universal DNA barcode for plants and animals. PLoS One 5:e13102PubMedCentralPubMedCrossRefGoogle Scholar
  18. 18.
    Li D-Z, Gao L-M, Li H-T, Wang H, Ge X-J, Liu J-Q, Chen Z-D, Zhou S-L, Chen S-L, Yang J-B, Fu C-X, Zeng C-X, Yan H-F, Zhu Y-J, Sun Y-S, Chen S-Y, Zhao L, Wang K, Yang T, Duan G-W, China Plant BOL Group (2011) Comparative analysis of a large dataset indicates that internal transcribed spacer (ITS) should be incorporated into the core barcode for seed plants. PNAS 108:19641–19646PubMedCrossRefGoogle Scholar
  19. 19.
    Hollingsworth PM (2011) Refining the DNA barcode for land plants. PNAS 108:19451–19452PubMedCrossRefGoogle Scholar
  20. 20.
    Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739PubMedCrossRefGoogle Scholar
  21. 21.
    White T, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis M, Gelfand D, Sninsky J, White T (eds) PCR protocols: a guide to methods and applications. Academic, New York, USA, pp 315–322Google Scholar
  22. 22.
    Sun Y, Skinner D, Liang G, Hulbert S (1994) Phylogenetic analysis of Sorghum and related taxa using internal transcribed spacers of nuclear ribosomal DNA. Theor Appl Genet 89:26–32PubMedCrossRefGoogle Scholar

Copyright information

© Springer New York 2014

Authors and Affiliations

  • Pascale Besse
    • 1
  1. 1.UMR C53 PVBMT Université de la Réunion – CiradUniversité de la RéunionIle de la RéunionFrance

Personalised recommendations