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Liquid medium and liquid overlays improve embryogenic tissue initiation in conifers

  • Cell Biology and Morphogenesis
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Abstract

Somatic embryogenesis (SE) is expected to play an important role in increasing productivity, sustainability, and uniformity of future US forests. For commercial use, SE technology must work with a variety of genetically diverse trees. Initiation in loblolly pine (Pinus taeda L.), the main commercial US forest species, is often recalcitrant for desirable genotypes. Liquid initiation medium with no or low gelling agent or placement of the explant on gelled medium followed later by a liquid medium overlay during the initiation process increased initiation for loblolly pine and Norway spruce (Picea abies). Loblolly pine liquid medium required reduction of NAA from 2 mg/l in gelled medium to 0.3 mg/l in liquid medium. Once the NAA concentration was adjusted, loblolly pine initiation occurred in liquid medium with fully immersed megagametophytes, explants supported at the liquid medium surface, or on gelled medium overlaid with liquid medium. Liquid overlays (0.25 ml) consisting of medium with NAA reduced to 0.3 mg/l, 9 mg/l ABA and no gelling agent applied to explants on 2 ml of gelled medium provided excellent initiation results. Greatest initiation percentages occurred when the liquid overlay was applied 14 days after placement of the megagametophyte on gelled medium. Initiation increases ranged from +8.5% with high-value cross-pollinated seed sources to +6.5 to +9.9% with open-pollinated and often recalcitrant seed sources. Liquid medium addition allows rapid replenishment of nutrients and adjustment or change of pH, hormones, or other parameters without disturbing the tissue.

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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

LP:

Loblolly pine

MES:

2(N-morpholino)ethanesulphonic acid buffer agent

NAA:

α-Naphthalene acetic acid

NS:

Norway spruce

OP:

Open-pollinated

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Acknowledgments

We thank the member companies of IPST and IPST at Georgia Tech for financial support and Bowater Inc., The Timber Company, Westvaco Corp. and MeadWestvaco Corp., Temple Inland, Texas Forest Service, and Weyerhaeuser Company for cones. We are grateful for the help of J. Dregar, S. Duncan, J. Grabowski, R. Gupta, J. Halpin, R. Howie, C. Umejiego, and Y. Zhang.

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

    Gerald S. Pullman & Anna Skryabina

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  1. Gerald S. Pullman
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  2. Anna Skryabina
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Correspondence to Gerald S. Pullman.

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Communicated by G.C. Phillips.

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Pullman, G.S., Skryabina, A. Liquid medium and liquid overlays improve embryogenic tissue initiation in conifers. Plant Cell Rep 26, 873–887 (2007). https://doi.org/10.1007/s00299-006-0296-1

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  • DOI: https://doi.org/10.1007/s00299-006-0296-1

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