Journal of Forestry Research

, Volume 24, Issue 2, pp 227–236 | Cite as

Analysis of genetic relationship among Arbutus unedo L. genotypes using RAPD and SSR markers

  • Filomena GomesEmail author
  • Rita Costa
  • Maria M. Ribeiro
  • Elisa Figueiredo
  • Jorge M. Canhoto
Original Paper


The strawberry tree (Arbutus unedo L.) is an underutilized, drought tolerant, fire resistant species with a south western distribution in Europe, and with ecological and putative socio-economical impact in Portugal and Mediterranean countries. Our aim was to develop an appropriate set of molecular markers to enable genetic diversity to be assessed and to fingerprint Arbutus unedo genotypes for breeding and conservation purposes in Portugal. Twenty-seven trees from a broad geographic range were screened with 20 random amplified polymorphic DNA (RAPD primers) and 11 microsatellite markers (SSR). The RAPDs generated 124 bands, 57.3% of which were polymorphic, with an expected heterozygosity of 27%. We cross-amplified 11 SSR primers developed for Vaccinium spp., and 5 were found to be polymorphic in A. unedo, with 75% of expected heterozygosity, a number of alleles of 11.6, a null allele frequency of 7.6% and a polymorphic information content of 71%. Although the SSRs were more polymorphic and informative than the RAPDs, both markers displayed high genetic variability with the gathered data. No geographic pattern was observed in the genetic variation distribution based on both marker systems, and the lack of correlation between genetic and geographical matrices was confirmed by Mantel tests. Likely, no correlation was found between pairwise SSR and RAPD band-sharing matrices. These results and their implications on A. unedo breeding and conservation programs are discussed.


Ericaceae fingerprinting geographic pattern molecular markers strawberry tree 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Agrama HA, Tuinstra MR. 2003. Phylogenetic diversity and relationships among sorghum accessions using SSRs and RAPDs. African Journal of Biotechnology, 2: 334–340.Google Scholar
  2. Albert T, Raspe O, Jacquemart AL. 2003. Clonal structure in Vaccinium myrtillus L. revealed by RAPD and AFLP markers. International Journal of Plant Sciences, 164: 649–655.CrossRefGoogle Scholar
  3. Albert T, Raspe O, Jacquemart AL. 2005. Diversity and spatial structure of clones in Vaccinium uliginosum populations. Canadian Journal of Botany, 83: 211–219.CrossRefGoogle Scholar
  4. Avramidou E, Ganopoulos IV, Aravanopoulos FA. 2010. DNA fingerprinting of elite Greek wild cherry. Forestry, 83: 527–533.CrossRefGoogle Scholar
  5. Badenes M, Garcés A, Romero C, Romero M, Clavé J, Rovira M, Llácer G. 2003. Genetic diversity of introduced and local Spanish persimmon cultivars revealed by RAPD markers. Genetic Resources and Crop Evolution, 50: 579–585.CrossRefGoogle Scholar
  6. Bassil NV, Boches P, Rowland L. 2006. Genetic diversity in the highbush blueberry evaluated with microsatellite markers. Journal of the American Society for Horticultural Science, 131: 674–686.Google Scholar
  7. Bassil NV, Bunch T, Nyberg A, Hummer K, Zee FT. 2010. Microsatellite markers distinguish Hawailan ohelo from other Vaccinium L. section Myrtillus species. Acta Horticulturae, 859: 81–89.Google Scholar
  8. Beland JD, Krakowski J, Ritland CE, Ritland K, El-kassaby YA. 2005. Genetic structure and mating system of northern Arbutus menziesii (Ericaceae) populations. Canadian Journal of Botany, 83: 1581–1589.CrossRefGoogle Scholar
  9. Boches PS, Bassil NV, Rowland LJ. 2005. Microsatellite markers for Vaccinium from EST and genomic library. Molecular Ecology, 5: 657–660.CrossRefGoogle Scholar
  10. Botstein WRL, Skolnick K, Davis RW. 1980. Construction of a genetic linkage map in man using restriction fragment length polymorphism. American Journal of Human Genetics, 32: 314–331.PubMedGoogle Scholar
  11. Celikel G, Demirsoy L, Demirsoy H. 2009. The strawberry tree (Arbutus unedo L.) selection in Turkey. Scientia Horticulturae, 118: 115–119.CrossRefGoogle Scholar
  12. Debnath SC. 2007. An assessment of the genetic diversity within a collection of wild Cranberry (Vaccinium macrocarpon Ait.) clones with RAPD-PCR. Genetic Resources and Crop Evolution, 54: 509–517.CrossRefGoogle Scholar
  13. Debnath SC. 2010. Berry bioreactor micropropagation and molecular analysis for genetic fidelity. Acta Horticulturae, 865: 43–51.Google Scholar
  14. Derory J, Mariette S, Gonzaléz-Martínez SC, Chagné D, Madur D, Gerber S, Brach J, Persyn F, Ribeiro MM, Plomion C. 2002. What can nuclear microsatellites tell us about maritime pine genetic resources conservation and provenance certification strategies? Annals of Forest Science, 59: 699–709.CrossRefGoogle Scholar
  15. Dramstad W. 1996. Do bumblebees (Hymenoptera: Apidae) really forage close to their nests? Journal of Insect Behavior, 9: 163–182.CrossRefGoogle Scholar
  16. El-Mousadik A, Petit RJ. 1996. Chloroplast DNA phylogeography of the argan tree of Morocco. Molecular Ecology, 5: 547–555.PubMedCrossRefGoogle Scholar
  17. Ellegren H. 2004. Microsatellites: simple sequences with complex evolution. Nature Reviews Genetics, 5: 435–445.PubMedCrossRefGoogle Scholar
  18. Fernandes L, Rocheta M, Cordeiro J, Pereira S, Gerber S, Oliveira, MM, Ribeiro MM. 2009. Genetic variation, matting patterns and gene flow in a Pinus pinaster Aiton clonal seed orchard. Annals of Forest Science, 65: 1–9.Google Scholar
  19. Godinho-Ferreira PG, Azevedo AM, Rego F. 2005. Carta da tipologia florestal de Portugal Continental. Silva Lusitana, 13: 1–34.Google Scholar
  20. Gomes F, Canhoto JM. 2009. Micropropagation of strawberry tree (Arbutus unedo L.) from adult plants. In Vitro Cellular & Developmental Biology — Plant, 45: 72–82.CrossRefGoogle Scholar
  21. Gomes F, Simões M, Lopes ML, Canhoto, JM. 2010. Effect of plant growth regulators and genotype on the micropropagation of adult trees of Arbutus unedo L. (strawberry tree). New Biotechnology, 27: 882–892.PubMedCrossRefGoogle Scholar
  22. Gupta PK, Varshney RK. 2000. The development and use of microsatellite markers for genetic analysis and plant breeding with special emphasis on bread wheat. Euphytica, 113: 163–185.CrossRefGoogle Scholar
  23. Hileman LC, Vasey MC, Thomas Parker V. 2001. Phylogeny and Biogeography of the Arbutoideae (Ericaceae): Implications for the Madrean-Tethyan Hypothesis. Systematic Botany, 26: 131–143.Google Scholar
  24. Hirai M, Yoshimura S, Ohsako T, Kubo N. 2010. Genetic diversity and phylogenetic relationships of the endangered species Vaccinium sieboldii and Vaccinium ciliatum (Ericaceae). Plant Systematics and Evolution, 287: 75–84.CrossRefGoogle Scholar
  25. Jordano P, García C, Godoy JA, García-Castaño JL. 2007. Differential contribution of frugivores to complex seed dispersal patterns. Proceedings National Academy of Sciences USA, 104: 3278–3282.CrossRefGoogle Scholar
  26. Kameyama Y, Kasagi T, Kudo, G. 2006. Eight microsatellite markers for sympatric alpine shrubs, Phyllodoce aleutica and P. caerulea (Ericaceae). Molecular Ecology, 6: 402–404.CrossRefGoogle Scholar
  27. Klooster MR, Hoenle AW, Culley TM. 2009. Characterization of microsatellite loci in the myco-heterotrophic plant Monotropa hypopitys (Ericaceae) and amplification in related taxa. Molecular Ecology Resources, 9: 219–221.PubMedCrossRefGoogle Scholar
  28. Kron KA, Stevens PF, Crayn DM, Anderberg AA, Gadek PA, Quinn CJ, Luteyn JL. 2002. Phylogenetic classification of Ericaceae: molecular and morphological evidence. The Botanical Review, 68: 335–423.CrossRefGoogle Scholar
  29. Lynch M. 1990. The similarity index and DNA fingerprinting. Molecular Biology Evolution, 7: 478–484.Google Scholar
  30. Majourhat K, Jabba Y, Hafidi A, Martínez-Gómez P. 2009. Molecular characterization and genetic relationships among most common identified morphotypes of critically endangered rare Moroccan species Argania spinosa (Sapotaceae) using RAPD and SSR markers. Annals of Forest Science, 65: 1–6.Google Scholar
  31. Mariette S, Chagne D, Decroocq S, Vendramin GG, Lalanne C, Madur D, Plomion C. 2001. Microsatellite markers for Pinus pinaster Ait. Annals of Forest Science, 58: 203–206.CrossRefGoogle Scholar
  32. Marshall TC, Slate J, Kruuk L, Pemberton JM. 1999. Statistical confidence for likelihood-based paternity inference in natural populations. Molecular Ecology, 7: 639–655.CrossRefGoogle Scholar
  33. Meireles C, Gonçalves P, Rego F, Silveira S. 2005. Estudo da regeneração natural das espécies arbóreas autóctones na Reserva Natural da Serra da Malcata. Silva Lusitana, 13: 217–231.Google Scholar
  34. Milne RI, Abbott RJ. 2009. Reproductive isolation among two interfertile Rhododendron species: low frequency of post-F-1 hybrid genotypes in alpine hybrid zones. Molecular Ecology, 17: 1108–1121.CrossRefGoogle Scholar
  35. Naval M, Zuriaga E, Pecchioli S, Llácer G, Giordani E, Badenes M. 2010. Analysis of genetic diversity among persimmon cultivars using microsatellite markers. Tree Genetics & Genomes, 6: 677–687.CrossRefGoogle Scholar
  36. Nei M. 1987. Molecular evolutionary genetics. New York: Columbia University Press, p.512.Google Scholar
  37. Pedro JG. 1994. Portugal Atlas do Ambiente. Notícia Explicativa II. 6 Carta da distribuição de figueira e medronheiro. Lisboa: Ministério do Ambiente e Recursos Naturais, p.39.Google Scholar
  38. Piotto B, Piccini C, Arcadu P. 2001. La ripresa della vegetazione dopo gli incendi nella regione mediterrânea. In: B. Piotto and A. Noi (eds.), Propagazione per seme di alberi e arbusti della flora mediterranea. Roma: Dipartimento Prevenzione e Risanamento Ambientali, pp. 32–39.Google Scholar
  39. Powell W, Morgante M, Andre C, Hanafey M, Vogel J, Tingey S, Rafalski A. 1996. The comparison of RFLP, RAPD, AFLP and SSR (microsatellite) markers for germplasm analysis. Molecular Breeding, 2: 225–239.CrossRefGoogle Scholar
  40. Rajora OP, Rahman MH. 2003. Microsatellite DNA and RAPD fingerprinting, identification and genetic relationships of hybrid poplar (Populus x canadensis) cultivars. Theoretical and Applied Genetics, 106: 470–477.PubMedGoogle Scholar
  41. Ribeiro MM, Plomion C, Petit R, Vendramin GG, Szmidt AE. 2001. Variation in chloroplast single-sequence repeats in Portuguese maritime pine (Pinus pinaster Ait.). Theoretical and Applied Genetics, 102: 97–103.CrossRefGoogle Scholar
  42. Rohlf FJ. 1997. NTSYS-pc. Numerical taxonomy and multivariate analysis system version 2.01. New York: Dep. of Ecology and Evolution, State University of New York, p.39.Google Scholar
  43. Schneller J, Holderegger R, Gugerli F, Eichenberger K, Lutz E. 1999. Patterns of genetic variation detected by RAPDs suggest a single origin with subsequent mutations and long-distance dispersal in the apomictic fern Dryopteris remota (Dryopteridaceae). American Journal of Botany, 85: 1038–1042.CrossRefGoogle Scholar
  44. Sokal R. 1979. Testing statistical significance of geographic variation patterns. Systematic Zoology, 28: 227–232.CrossRefGoogle Scholar
  45. Sorkheh K, Shiran B, Kiani S, Amirbakhtiar N, Mousavi S, Rouhi V, Mohammady-D S, Gradziel TM, Malysheva-Otto LV, Martínez-Gómez P. 2009. Discriminating ability of molecular markers and morphological characterization in the establishment of genetic relationships in cultivated genotypes of almond and related wild species. Journal Forestry Research, 20: 183–194.CrossRefGoogle Scholar
  46. Spooner D, Van Treuren R, Vicente MC. 2005. Molecular Markers for Genebank Management. Rome: International Plant Genetic Resources Institute (IPGRI), Technical Bulletin No 10.Google Scholar
  47. Takrouni MM, Boussaid M. 2010. Genetic diversity and population’s structure in Tunisian strawberry tree (Arbutus unedo L.). Scientia Horticulturae, 126: 330–337.CrossRefGoogle Scholar
  48. Tan X-X, Li Y, Ge X-J. 2009. Development and characterization of eight polymorphic microsatellites for Rhododendron simsii Planch (Ericaceae). Conservation Genetics, 10: 1553–1555.CrossRefGoogle Scholar
  49. Torres JA, Valle F, Pinto C, Garcia-Fuentes A, Salazar C, Cano E. 2002. Arbutus unedo communities in southern Iberian Peninsula mountains. Plant Ecology, 160: 207–223.CrossRefGoogle Scholar
  50. Vaughan SP, Cottrell JE, Moodley DJ, Connolly T, Russell1 K. 2007. Distribution and fine-scale spatial-genetic structure in British wild cherry (Prunus avium L.). Heredity, 98: 274–283.PubMedCrossRefGoogle Scholar
  51. Vekemans X, Beauwens T, Lemaire M, Roldan-Ruiz I. 2002. Data from amplified fragment polymorphism (AFLP) markers show indication of size homoplasy and of a relationship between degree of homoplasy and fragment size. Molecular Ecology, 11: 139–151.PubMedCrossRefGoogle Scholar
  52. Wang N, Qin ZC, Yang JB, Zhang JL. 2010. Development and characterization of 15 microsatellite loci for Rhododendron delavayi Franch. (Ericaceae). Hortscience, 45: 457–459.Google Scholar
  53. Weir CC, Cockerham BS. 1984. Estimating F-statistics for the analysis of population structure. Evolution, 38: 1358–1370.CrossRefGoogle Scholar
  54. Wohlgemuth T, Bürgi M, Scheidegger C, Schütz M. 2002. Dominance reduction of species through disturbance — a proposed management principle for Central European forests. Forest Ecology and Management: 166, 1–15.CrossRefGoogle Scholar

Copyright information

© Northeast Forestry University and Springer-Verlag Berlin Heidelberg 2012

Authors and Affiliations

  • Filomena Gomes
    • 1
    Email author
  • Rita Costa
    • 2
  • Maria M. Ribeiro
    • 3
  • Elisa Figueiredo
    • 4
  • Jorge M. Canhoto
    • 4
  1. 1.CERNAS, Departamento FlorestalEscola Superior Agrária CoimbraCoimbraPortugal
  2. 2.INRBInstituto Nacional de Recursos Biológicos, IP/L-INIAOeirasPortugal
  3. 3.Departamento de Recursos Naturais e Desenvolvimento SustentávelEscola Superior AgráriaCastelo BrancoPortugal
  4. 4.Centre of Functional Ecology, Department of Life SciencesUniversity of CoimbraCoimbraPortugal

Personalised recommendations