, Volume 85, Issue 1–3, pp 341–345 | Cite as

Monitoring genetic fidelity vs somaclonal variation in Norway spruce (Picea abies) somatic embryogenesis by RAPD analysis

  • Berthold Heinze
  • Josef Schmidt


Somaclonal variation, which is a welcome source of genetic variation for crop breeding, is unwanted when direct regenerants have to be used in tissue culture mass propagation (eg. in many forest trees), or in the regeneration of genetically transformed plants. Random amplified polymorphic DNA (RAPD) was used to analyse somatic embryos and plants regenerated from embryogenic cell lines in Norway spruce, Picea abies (L.) Karst. RAPD facilitated the identification of clones, as material from the same cell lines shared identical patterns of amplified fragments, whereas regenerants from different cell lines were easily distinguishable by their respective patterns. For comparisons with explant donor genotypes, cell lines were initiated from cotyledons. Some of the seedlings that had parts of their cotyledons removed were grown on as control plants. Somatic embryos regenerated from cotyledon cell lines showed no aberrations in RAPD banding patterns with respect to donor plants. We conclude that gross somaclonal variation is absent in our plant regeneration system.

Key words

Picea abies genetic stability somatic embryogenesis RAPD 



embryogenic suspensor mass


random amplified polymorphic DNA


restriction fragment length polymorphism

(2,4-dichlorophenoxy)acetic acid


1-naphthaleneacetic acid



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  1. Attree S.M. & L.C. Fowke, 1993. Embryogeny of gymnosperms: advances in synthetic seed technology of conifers. Plant Cell Tissue Organ Cult. 35: 1–35.CrossRefGoogle Scholar
  2. Bebeli P.J., A. Karp & P.J. Kaltsikes, 1990. Somaclonal variation from cultured immature embryos of sister lines of rye differing in heterochromatin content. Genome 33: 177–183.Google Scholar
  3. Becwar M.R., 1993. Conifer somatic embryogenesis and clonal forestry. p. 200–223. In: M.R. Ahuja & W.J. Libby (Eds). Clonal Forestry I. Springer, Berlin.Google Scholar
  4. Becwar M.R., T.L. Noland & S.R. Wann, 1987. Somatic embryo development and plant regeneration from embryogenic Norway spruce callus. TAPPI J. 70: 155–160.Google Scholar
  5. Breiman A., T. Felsenburg & E. Galun, 1987. Nor loci analysis in progenies of plants regenerated from the scutellar callus of bread-wheat. A molecular approach to evaluate somaclonal variation. Theor. Appl. Genet. 73: 827–831.CrossRefGoogle Scholar
  6. Brettell R.I.S., M.A. Pallotta, J.P. Gustafson & R. Appels, 1986. Variation at the Nor loci in triticale derived from tissue culture. Theor. Appl. Genet. 71: 637–643.CrossRefGoogle Scholar
  7. Brown P.T.H., 1991. The spectrum of molecular changes associated with somaclonal variation. IAPTC Newsletter 66: 14–25.Google Scholar
  8. Brown P.T.H., E. Goebel & H. Loerz, 1991. RFLP analysis of Zea mays callus cultures and their regenerated plants. Theor. Appl. Genet. 81: 227–232.CrossRefGoogle Scholar
  9. Butigereit, J., 1991. Somatische Embryogenese bei Picea abies (L.) Karst. Vienna, University of Vienna, Dissertation.Google Scholar
  10. Cactano-Anollés, G., B.J. Bassam & P.M. Gresshoff, 1993. Enhanced detection of polymorphic DNA. Mol. Gen. Genet. in press.Google Scholar
  11. Carlson J.E., L.K. Tulsieram, J.C. Glaubitz, V.W.K. Luk, C. Kauffeidt & R. Rutledge, 1991. Segregation of random amplified DNA markers in F1 progeny of conifers. Theor. Appl. Genet. 83: 194–200.CrossRefGoogle Scholar
  12. Eastman P.A.K., F.B. Webster, J.A. Pitel & D.R. Roberts, 1991. Evaluation of somaclonal variation during somatic embryogenesis of interior spruce (Picea glauca engelmanni complex) using culture morphology and isozyme analysis. Plant Cell Rep. 10: 425–430.CrossRefGoogle Scholar
  13. Gupta P.K., G. Pullman, R. Timmis, M. Kreitinger, W.C. Carlson, J. Grob & E. Welty, 1993. Forestry in the 21 st century. The biotechnology of somatic embryogenesis. Biotechnology 11: 454–459.CrossRefGoogle Scholar
  14. Hakman I., L.C. Fowke, S.von Arnold & T. Eriksson, 1985. The development of somatic embryos in tissue cultures initiated from immature embryos of Picea abies (Norway spruce). Plant Sci. 38: 53–59.CrossRefGoogle Scholar
  15. Harada T., K. Matsukawa, T. Sato, R. Ishikawa, M. Niizeki & K. Saito, 1993. DNA-RAPDs detect genetic variation and patemity in Malus. Euphytica 65: 87–91.CrossRefGoogle Scholar
  16. Heinze, B., 1993. Genetic stability in Norway spruce somatic embryogenesis as analysed by randomly amplified DNA. Vienna, Universität für Bodenkultur, Dissertation.Google Scholar
  17. Heinze B., 1994. RAPD reactions from crude plant DNA: Adding RNase A as a ‘helper enzyme’. Mol. Biotechnol. 1: 307–310.PubMedCrossRefGoogle Scholar
  18. Isaoel N., L. Tremblay, M. Michaud, F.M. Tremblay & J. Bousquet, 1993. RAPDs as an aid to evaluate the genetic integrity of somatic embryogenesis-derived populations of Picea mariana (Mill). BSP. Theor. Appl. Genet. 86: 81–87.Google Scholar
  19. Klein-Lankhorst R.M., A. Vermunt, R. Weide, T. Liharska & P. Zabel, 1991. Isolation of molecular markers for tomato (L. escuientum) using random amplified polymorphic DNA (RAPD). Theor. Appl. Genet. 83: 108–114.CrossRefGoogle Scholar
  20. Krogstrup P., 1986. Embryolike structures from cotyledons and ripe embryos of Norway spruce (Picea abies). Can. J. For. Res. 16: 664–668.CrossRefGoogle Scholar
  21. Larkin P.J. & W.R. Scowcroft, 1981. Somaclonal variation—a novel source of variability from cell cultures for plant improvement. Theor. Appl. Genet. 60: 197–214.CrossRefGoogle Scholar
  22. Lelu M.-A., 1987. Variations morphologiques et genetiques chez Picea abies obtenues apres embryogenese somatique. Ann. AFOCEL 1987: 35–47.Google Scholar
  23. Lelu, M.A., M. Boulay & Y. Arnaud, 1987. Obtention de cals embryogenes a partir de cotyledons de Picea abies (L.) Karst. preleves sur de jeunes plantes agees de 3 a 7 jours apres germination. C.R. Acad. Sci. Paris t. 305 Serie III: 105–109.Google Scholar
  24. Mo L.H., S.von Arnold & U. Lagercrantz, 1989. Morphogenic and genetic stability in longterm embryogenic cultures and somatic embryos of Norway spruce (Picea abies (L.) Karst.). Plant Cell Rep. 8: 375–378.CrossRefGoogle Scholar
  25. Mulcahy D.L., M. Cresti, S. Sansavini, G.C. Douglas, H.F. Linskens, G.B. Mulcahy, R. Vignani & M. Pancaldi, 1993. The use of random amplified polymorphic DNAs to fingerprint apple genotypes. Sci. Hort. Amsterdam 54: 89–96.CrossRefGoogle Scholar
  26. Pikaart M.J. & B. Villeponteau, 1993. Suppression of PCR amplification by high levels of RNA. Bio Techniques 14: 24–27.Google Scholar
  27. Rafalski A., S. Tingey & J.G.K. Williams, 1993. Random amplified polymorphic DNA (RAPD) markers. p. 1–9. In: S.B. Gelvin, R.A. Schilperoort & D.P.S. Verma (Eds). Plant Molecular Biology Manual. Kluwer, Dordrecht.Google Scholar
  28. Roberts D.R., F.B. Webster, B.S. Flinn, W.R. Lazaroff & D.R. Cyr, 1993. Somatic embryogenesis of spruce. p. 427–450. In: K. Redenbaugh (Ed). Synseeds. CRC Press, Boca Raton.Google Scholar
  29. Roberts D.R., B.C.S. Sutton & B.S. Flinn, 1990. Synchronous and high frequency germination of interior spruce somatic embryos following partial drying at high relative humidity. Can. J. Bot. 68: 1086–1090.Google Scholar
  30. Shenoy V.B. & I.K. Vasil, 1992. Biochemical and molecular analysis of plants derived from embryogenic tissue cultures of napier grass (Pennisetum purpureum K. Schum). Theor. Appl. Genet. 83: 947–955.CrossRefGoogle Scholar
  31. Son S.H., H.K. Moon & R.B. Hall, 1993. Somaclonal variation in plants regenerated from callus culture of hybrid aspen (Populus alba L. × P. grandidentata Michx). Plant Sci. 90: 89–94.CrossRefGoogle Scholar
  32. Stiles J.I., C. Lemme, S. Sondur, M.B. Morshidi & R. Manshardt, 1993. Using randomly amplified polymorphic DNA for evaluating genetic relationships among papaya cultivars. Theor. Appl. Genet. 85: 697–701.CrossRefGoogle Scholar
  33. Torres A.M., T. Millan & J.I. Cubero, 1993. Identifying rose cultivars using random amplified polymorphic DNA markers. HortScience 28: 333–334.Google Scholar
  34. Tulsieram L.K., J.C. Glaubitz, G. Kiss & J.E. Carlson, 1992. Single tree genetic linkage mapping in conifers using haploid DNA from megagametophytes. Biotechnology 10: 686–690.PubMedCrossRefGoogle Scholar
  35. Vierling R.A. & H.T. Nguygen, 1992. Use of RAPD markers to determine the genetic diversity of diploid, wheat genotypes. Theor. Appl. Genet. 84: 835–838.CrossRefGoogle Scholar
  36. von Arnold S. & I. Hakman, 1988. Regulation of somatic embryo development in Picea abies by abscisic acid (ABA). J. Plant Physiol. 132: 164–169.Google Scholar
  37. Welsh J. & M. McClelland, 1990. Fingerprinting genomes using PCR with arbitrary primers. Nucl. Acid. Res. 18: 7213–7218.CrossRefGoogle Scholar
  38. Wilde J., R. Waugh & W. Powell, 1992. Genetic fingerprinting of Theobroma clones using randomly amplified polymorphic DNA markers. Theor. Appl. Genet. 83: 871–877.CrossRefGoogle Scholar
  39. Williams C.E. & D.A.St. Clair, 1993. Phenetic relationships and levels of variability detected by restriction fragment length polymorphism and random amplified polymorphic DNA analysis of cultivated and wild accessions of Lycopersicon esculentum. Genome 36: 619–630.PubMedCrossRefGoogle Scholar
  40. Williams J.G.K., A.R. Kubelik, K.J. Livak, J.A. Rafalski & S.V. Tingey, 1990. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucl. Acid. Res. 18: 6531–6535.CrossRefGoogle Scholar

Copyright information

© Kluwer Academic Publishers 1995

Authors and Affiliations

  • Berthold Heinze
    • 1
  • Josef Schmidt
    • 1
  1. 1.Biotechnology DepartmentAustrian Research CentreSeibersdorfAustria

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