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Gibberellin inhibitors improve embryogenic tissue initiation in conifers

  • Cell Biology and Morphogenesis
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Abstract

Somatic embryogenesis (SE), the most promising technology to multiply high-value coniferous trees from advanced breeding and genetic engineering programs, is expected to play an important role in increasing productivity, sustainability, and uniformity of future forests in the United States. For commercial use, SE technology must work with a variety of genetically diverse trees. Initiation in loblolly pine (LP; Pinus taeda L.), our main focus species, is often recalcitrant for desirable genotypes. Initiation of LP, slash pine (SP; Pinus elliottii), Douglas-fir (DF; Pseudotsuga menziesii), and Norway spruce (NS; Picea abies) were improved through the use of paclobutrazol, a gibberellin synthesis inhibitor. Paclobutrazol was effective at concentrations ranging from 0.25 mg/l to 3.0 mg/l (0.85–10.2 μM) and optimal in LP at 1.0 mg/l. Using control media (no paclobutrazol) and 0.33–1.0 mg/l paclobutrazol, initiation percentages in LP, SP, DF, and NS were improved from 37.7% to 44.2% (across experiments), 19.3% to 28.5%, 16.9% to 23.7%, and 38.8% to 48.5%, respectively. Other gibberellin inhibitors such as flurprimidol, chlormequat-Cl, and daminozide also caused statistically significant increases in LP initiation when added to the medium at concentrations of 0.34, 10.0, and 1.0 μM, respectively. No detrimental effects on subsequent embryo development were observed when 29 new initiations from medium without GA inhibitor and 28 new initiations from medium containing paclobutrazol were tracked through culture capture, liquid culture establishment, cotyledonary embryo development, and germination.

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Abbreviations

ABA:

Abscisic acid

AC:

Activated carbon

BA:

6-Benzylaminopurine

8-Br-cGMP:

Guanosine 3′,5′-cyclic monophosphate, 8-bromo-, sodium salt

2,4-D:

2,4-Dichlorophenoxyacetic acid

DF:

Douglas-fir

GA:

Gibberellin

LP:

Loblolly pine

MES:

2(N-Morpholino)ethanesulphonic acid buffer agent

NAA:

α-Naphthalene acetic acid

NS:

Norway spruce

OP:

Open-pollinated

SP:

Slash pine

References

  • Biddington NL, Sutherland RA, Robinson HT (1992) The effects of gibberellic acid, fluridone, abscisic acid, and paclobutrazol on anther culture of brussels sprouts. Plant Growth Regul 11:81–84

    CAS  Google Scholar 

  • Burkhart LF, Meyer MM Jr (1991) The gibberellin synthesis inhibitors, ancymidol and flurprimidol, promote in vitro rooting of white pine microshoots. Plant Cell Rep 10:475–476

    Article  CAS  Google Scholar 

  • Buta JG, Spaulding DW (1991) Effect of paclobutrazol on abscisic acid levels in wheat seedlings. J Plant Growth Regul 10:59–61

    CAS  Google Scholar 

  • Forest Service FIA Data Retrieval System (2004) http://ncrs2.fs.fed.us/4801/fiadb/index.htm (courtesy of A. Goetzl, AFPA)

  • Gmitter FG Jr, Moore GA (1986) Plant regeneration from undeveloped ovules and embryogenic calli of Citrus: embryo production, germination and plant survival. Plant Cell Tissue Organ Cult 6:139–147

    Article  CAS  Google Scholar 

  • Gomez-Cadenas A, Aentella R, Walker-Simmons MK, Ho T-HD (2001) Gibberellin/abscisic acid antagonism in barley aleurone cells: site of action of the protein kinase PKABA1 in relation to gibberellin signaling molecules. Plant Cell 13:667–679

    Article  CAS  PubMed  Google Scholar 

  • Graham JS, Hobbs SD, Zaerr JB (1994) The effect of flurprimidol on bud flush, shoot growth, and on endogenous gibberellins and abscisic acid on Douglas-fir seedlings. J Plant Growth Regul 13:131–136

    Article  CAS  Google Scholar 

  • Halperin W (1970) Embryos from somatic plant cells. In: Padykula HA (ed) Control mechanisms in the expression of cellular phenotypes (Symp Int Soc for Cell Biol). Academic, New York, London, pp 169–191

    Google Scholar 

  • Hare RC (1984) EL-500: an effective growth retardant for dwarfing southern pine seedlings. Can J For Res 14:123–127

    CAS  Google Scholar 

  • Hita O, Lafarga C, Guerra H (1997) Somatic embryogenesis from chickpea (Cicer areitinum L.) immature cotyledons: the effect of zeatin, gibberellic acid, and indole-3-butyric acid. Acta Physiol Plant 19:333–338

    CAS  Google Scholar 

  • Hutchinson MJ, KrishnaRaj S, Saxena PK (1997) Inhibitory effect of GA3 on the development of thidiazuron-induced somatic embryogenesis in geranium (Pelargonium × hortorum Bailey) hypocotyl cultures. Plant Cell Rep 16:435–438

    Article  CAS  Google Scholar 

  • Kong L, Attree SM, Fowke LC (1997) Changes of endogenous hormone levels in developing seeds, zygotic embryos and megagametophytes in Picea glauca. Physiol Plant 101:23–30

    Article  CAS  Google Scholar 

  • Li B, Wolyn DJ (1995) The effects of ancymidol, abscisic acid, uniconazole, and paclobutrazol on somatic embryogenesis of asparagus. Plant Cell Rep 14:529–533

    Article  CAS  Google Scholar 

  • Li B, Wolyn DJ (1997) Interaction of ancymidol with sucrose and α-napthaleneacetic acid in promoting asparagus (Asparagus officinalis L.) somatic embryogenesis. Plant Cell Rep 16:879–883

    Article  CAS  Google Scholar 

  • Liao YK, Amerson HV (1995) Slash pine (Pinus elliottii Engelm.) somatic embryogenesis I. Initiation of embryogenic cultures from immature zygotic embryos. New For 10:145–163

    Google Scholar 

  • Noma M, Huber J, Ernst D, Pharis R (1982) Quantitation of gibberellins and the metabolism of [3H] gibberellin A1 during somatic embryogenesis in carrot and anise cell cultures. Planta 155:369–376

    CAS  Google Scholar 

  • Prestemon JP, Abt RC (2002) Timber products supply and demand. In: Wear DN, Greis JG (eds) Southern forest resource assessment. Gen Tech Rep SRS-53. US Department of Agriculture, Forest Service, Southern Research Station, Asheville, pp 299–325

    Google Scholar 

  • Pullman GS (2003) Gibberellin synthesis inhibitors improve conifer embryogenic tissue initiation. 2003 congress on in vitro biology, 31 May–4 June 2003, Portland, Ore. In vitro 39: Abstract Spring 2003, P1039 p 28A. ISSN 1071-2690

    Google Scholar 

  • Pullman GS, Gupta PK (1994) Method for reproducing conifers by somatic embryogenesis using mixed growth hormones for embryo culture. US Patent 5 294 549. Issued 15 March 1994

  • Pullman G, Peter G (2002) Methods of initiating and proliferating embryogenic cultures in conifers. US patent application filed 19 February 2002, pending

  • Pullman GS, Webb DT (1994) An embryo staging system for comparison of zygotic and somatic embryo development. TAPPI R&D division biological sciences symposium, 3–6 October 1994, Minneapolis, Minn., pp 31–34

  • Pullman GS, Namjoshi K, Zhang Y (2003a) Somatic embryogenesis in loblolly pine (Pinus taeda L.): improving culture initiation with abscisic acid, silver nitrate, and cytokinin adjustments. Plant Cell Rep 22:85–95

    Article  CAS  PubMed  Google Scholar 

  • Pullman GS, Zhang Y, Phan B (2003b) Brassinolide improves embryogenic tissue initiation in conifers and rice. Plant Cell Rep 22:96–104

    Article  CAS  PubMed  Google Scholar 

  • Pullman GS, Montello P, Cairney J, Xu N, Feng X (2003c) Loblolly pine (Pinus taeda L.) somatic embryogenesis: maturation improvements by metal analyses of zygotic and somatic embryos. Plant Sci 164:955–969

    Article  CAS  Google Scholar 

  • Pullman GS, Johnson S, Van Tassel S, Zhang Y (2004) Somatic embryogenesis in loblolly pine (Pinus taeda L.): improving culture initiation with MES pH buffer, biotin, and folic acid. Plant Cell Tissue Organ Cult (in press)

  • Rademacher W (2000) Growth retardants: effects on gibberellin biosynthesis and other metabolic pathways. Annu Rev Plant Physiol Plant Mol Biol 51:501–531

    Article  CAS  PubMed  Google Scholar 

  • Rajasekaran K, Hein MB, Vasil IK (1987) Endogenous abscisic acid and indole-3-acetic acid and somatic embryogenesis in cultured leaf explants of Pennisetum purpureum Schum. Plant Physiol 84:47–51

    CAS  Google Scholar 

  • Rudus I, Kepczynska E, Kepczynski J (2000) Regulation of Medicago sativa L. somatic embryogenesis by gibberellins. Plant Growth Regul 36:91–95

    Article  Google Scholar 

  • Sankhla A, Davis TD, Sankhla D, Sankhla N, Upadhyaya A, Joshi S (1992) Influence of growth regulators on somatic embryogenesis, plant regulation, and post-transplant survival of Echinochloa frumentacea. Plant Cell Rep 11:368–371

    Article  CAS  Google Scholar 

  • Schultz RP (1999) Loblolly—the pine for the twenty-first century. New For 17:71–88

    Google Scholar 

  • Shimizu K, Nagaike H, Yabuya T, Adachi T (1997) Plant regeneration from suspension culture of Iris germanica. Plant Cell Tissue Organ Cult 50:27–31

    Article  Google Scholar 

  • Verhagen SA, Wann SR (1989) Norway spruce somatic embryogenesis: high-frequency initiation from light-cultured mature embryos. Plant Cell Tissue Organ Cult 16:103–111

    Article  Google Scholar 

  • White CN, Proesbsting WM, Hedden P, Rivin CJ (2000) Gibberellins and seed development in Maize. I. Evidence that gibberellin/abscisic acid balance governs germination versus maturation pathways. Plant Physiol 122:1081–1088

    Article  CAS  PubMed  Google Scholar 

  • Ziv M, Kahany S, Lilien-Kipnis H (1994) Scaled-up proliferation and regeneration of Nerine in liquid cultures. Part I. The induction and maintenance of proliferating meristematic clusters by paclobutrazol in bioreactors. Plant Cell Tissue Organ Cult 39:109–115

    CAS  Google Scholar 

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Acknowledgements

We thank the member companies of IPST for financial support and Bowater, Inc., International Paper Company, Rayonier, The Timber Company, Westvaco (now MeadWestvaco Corp.), and Weyerhaeuser Company for cones. We are grateful for the help of R. Chopra, J. Cordero, R. Gupta, J. Halpin, R. Howie, L. Lockhart, and C. Perfetti.

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Authors and Affiliations

  1. Institute of Paper Science and Technology, Georgia Institute of Technology, 500 10th Street NW, Atlanta, GA, 30332-0620, USA

    Gerald S. Pullman, J. Mein, S. Johnson & Y. Zhang

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  1. Gerald S. Pullman
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  2. J. Mein
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  3. S. Johnson
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  4. Y. Zhang
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Correspondence to Gerald S. Pullman.

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Communicated by G. Phillips

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Pullman, G.S., Mein, J., Johnson, S. et al. Gibberellin inhibitors improve embryogenic tissue initiation in conifers. Plant Cell Rep 23, 596–605 (2005). https://doi.org/10.1007/s00299-004-0880-1

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  • DOI: https://doi.org/10.1007/s00299-004-0880-1

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