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Factors affecting recurrent shoot multiplication in in vitro cultures of 17- to 20-year-old douglas fir trees


The effects of sucrose concentration, addition of ammonium nitrate, and exposure to N6-benzyl-adenine (BA) on multiplication potential with shoots derived from shoot cultures of 17- to 20-yr-old Douglas fir trees [Pseudotsuga menziesii (Mirb.) Franco] were compared. Each of these conditions, when compared independently, affected recurrent shoot multiplication and influenced shoot development, as measured by the abundance of shoot apices. Sucrose concentration was influential, the use of 25 g · liter−1 providing twice the multiplication obtained with 20 g · liter−1, and 14 × that obtained with the 30 g · liter−1 concentration routinely used (tree 11). Ammonium nitrate usage also improved multiplication, a 2.5 times improvement being obtained after incorporation of 100 mg · liter−1 NH4NO3 into the medium (tree 33). Shoot cultures were responsive but relatively sensitive to addition of BA, the best improvement in multiplication (5 times) being obtained with brief exposures to 3 mg · liter−1 BA (tree 11). Although shoot cultures were responsive to the conditions investigated, differences in shoot multiplication and development were not displayed for several weeks. It was not possible therefore to repeat all the treatments with more than one genotype; however, when this was possible a genotype-dependent variation in response was evident.

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  1. Ball, E. A. Tissue culture multiplication ofSequoia. In: Bonga, J. M.; Durzan, D. J., eds. Cell and tissue culture in forestry, vol 3. Dordrecht, The Netherlands: Martinus Nijhoff; 1987:146–158.

    Google Scholar 

  2. Bonga, J. M. Vegatative propagation in relation to juvenility, maturity, and rejuvenation. In: Bonga, J. M.; Durzan, D. J., eds. Tissue culture in forestry. The Hague, The Netherlands: Nijhoff/Junk; 1982:387–412.

    Google Scholar 

  3. Boulay, M. Propagationin vitro du Douglas (Pseudotsuga menziesii (Mirb Franco) par micropropagation de germination aseptique et culture de bourgeons dormants. In: Micropropagation d’arbres forestiers. Nangis, France: AFOCEL, Etudes et Recherches. 12:49–55; 1979.

    Google Scholar 

  4. Dunstan, D. I.; Mohammed, G. H.; Thorpe, T. A. Shoot production and elongation on explants from vegetative buds excised from 17 to 20-year-oldPseudotsuga menziesii. N. Z. J. For. Sci. 16:269–282; 1986.

    Google Scholar 

  5. Dunstan, D. I.; Bethune, T.; Kikcio, S., et al.In vitro shoot formation among 17-year-old Douglas fir provenances. Can. J. For. Res. 19:1299–1302; 1989.

    Google Scholar 

  6. Franclet, A.; David, A.; David, H., et al. Première mise en évidence morphologique d’un rajeunissement de méristèmes primaires caulinaires de Pin maritime âgé (Pinus pinaster Sol.). C. R. Acad. Sci. Ser. 3. 209:927–929; 1980.

    Google Scholar 

  7. Gupta, P. K.; Durzan, D. J. Micropropagation and phase specificity in mature, elite Douglas fir. J. Am. Soc. Hort. Sci. 112:969–971; 1987.

    Google Scholar 

  8. Hasnain, S.; Cheliak, W. Tissue culture in forestry: economic and genetic potential. For. Chron. 82:219–225; 1986.

    Google Scholar 

  9. Horgan, K.Pinus radiata. In: Bonga, J. M.; Durzan, D. J., eds.. Cell and tissue culture in forestry, vol 3. Dordrecht, The Netherlands: Martinus Nijhoff; 1987:128–145.

    Google Scholar 

  10. Krizek, D. T. Guidelines for measuring and reporting environmental conditions in controlled environment studies. Physiol. Plant. 56:231–235; 1982.

    Article  Google Scholar 

  11. Langford, P. J.; Wainwright, H. Effects of sucrose concentration on the photosynthetic ability of rose shootsin vitro. Ann. Bot. 60:633–640; 1987.

    CAS  Google Scholar 

  12. Lee, M. S.; Kirby, E. G. Growth parameters of cell suspension cultures ofPseudotsuga menziesii and effects of nitrogen sources on growth. N. Z. J. For. Sci. 16:369–376; 1986.

    Google Scholar 

  13. Mohammed, G. H.; Patel, K. R.; Vidaver, W. E. The control of adventitious root production in tissue-cultured Douglas fir. Can. J. For. Res. 19:1322–1329; 1989.

    CAS  Google Scholar 

  14. Montain, C. R.; Haissig, B. E.; Curtis, J. D. Differentiation of adventitious root primordia in callus ofPinus banksiana seedling cuttings. Can. J. For. Res. 13:195–200; 1983.

    Google Scholar 

  15. Pate, J. S.; Layzell, D. B. Energetics and biological costs of nitrogen assimilation. In: Miflin, B. J.; Lea, P. J., et al. The biochemistry of plants, vol 16. Intermediary nitrogen metabolism. NY: Academic Press; 1990:1–42.

    Google Scholar 

  16. Teasdale, R. D. Generation of sustainablePinus radiata cell suspension culture and studies of cellular nitrogen nutrition. N. Z. J. For. Sci. 16:377–386; 1986.

    Google Scholar 

  17. Thorpe, T. A.; Bagh, K.; Cutler, A. J., et al. A14N and15N nuclear magnetic resonance study of nitrogen metabolism in shoot-forming cultures of white spruce (Picea glauca) buds. Plant Physiol. 91:193–202; 1989.

    Article  PubMed  CAS  Google Scholar 

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Dunstan, D.I., Lashta, D.P., Kikcio, S.I. et al. Factors affecting recurrent shoot multiplication in in vitro cultures of 17- to 20-year-old douglas fir trees. In Vitro Cell Dev Biol - Plant 28, 33–38 (1992).

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