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Agrobacterium-mediated transformation of hybrid poplar suspension cultures and regeneration of transformed plants

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Abstract

A method for Agrobacterium-mediated transformation of hybrid poplar (Populus alba x P. grandidentata cv. ‘Crandon’) suspension cultures and regeneration of transformed plants is described. Transformants were recovered when suspension cultures were inoculated with Agrobacterium tumefaciens at a density of 107 colony-forming units ml-1, cocultivated for 48 h, and plated to cellulose acetate filters on Woody Plant Medium containing 4.5 μM 2,4-dichlorophenoxyacetic acid and 250 mg l-1 cefotaxime. Levels of cefotaxime greater than 250 mg l-1 were unnecessary for control of residual bacteria and inhibited callus growth. Transgenic plants were regenerated by culturing the transformed callus on media containing 0.11 to 27 μM thidiazuron. In contrast to thidiazuron, N6-benzyladenine had a negative effect on shoot regeneration; the callus became necrotic when we attempted to induce shoots with concentrations of 1.1 to 8.9 μM, and growth was inhibited when concentrations of 0.11 or 0.22 μM were used to regenerate callus from suspension cultures. Following cocultivation of poplar suspension cultures, we recovered transgenic plants containing the maize transposon Ac, and callus containing an insect toxin gene from Bacillus thuringiensis.

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Abbreviations

BA:

N6-benzyladenine

CIM:

callus initiation medium

CaMV:

cauliflower mosaic virus

cfu's:

colony-forming units

HPT:

hygromycin phosphotransferase

MS:

Murashige and Skoog medium (Murashige & Skoog 1962)

NPT-II:

neomycin phosphotransferase-II

PAR:

photosynthetically active radiation

PCR:

polymerase-chain-reaction

TDZ:

thidiazuron

WPM:

Woody Plant Medium (Lloyd & McCown 1980)

2,4-d :

2,4-dichlorophenoxyacetic acid

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Author notes

  1. Glenn T. Howe

    Present address: Department of Forest Resources, University of Minnesota, 55108, St. Paul, MN, USA

  2. Barry Goldfarb

    Present address: Department of Forestry, North Carolina State University, 27695, Raleigh, NC, USA

Authors and Affiliations

  1. Department of Forest Science, Oregon State University, 97331, Corvallis, OR, USA

    Glenn T. Howe, Barry Goldfarb & Steven H. Strauss

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  1. Glenn T. Howe
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  2. Barry Goldfarb
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  3. Steven H. Strauss
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Howe, G.T., Goldfarb, B. & Strauss, S.H. Agrobacterium-mediated transformation of hybrid poplar suspension cultures and regeneration of transformed plants. Plant Cell Tiss Organ Cult 36, 59–71 (1994). https://doi.org/10.1007/BF00048316

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  • DOI: https://doi.org/10.1007/BF00048316

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