An evaluation of antibiotics for the elimination of Agrobacterium tumefaciens from walnut somatic embryos and for the effects on the proliferation of somatic embryos and regeneration of transgenic plants |
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Authors: | H Tang Z Ren G Krczal |
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Institution: | (1) Zentrum für Grüne Gentechnik, Staatliche Lehr- und Forschungsanstalt für Landwirtschaft, Weinbau und Gartenbau, 67435 Neustadt an der Weinstrasse, Germany Fax: 06321-671222 e-mail: gkrczal.slfa-nw@agrarinfo.rpl.de, DE |
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Abstract: | Four antibiotics were evaluated for their effects on eliminating the hypervirulent Agrobacterium tumefaciens strain C58C1 ATHV RifR (pEHA101)/p35-gus-intron from walnut somatic embryos and on the production of secondary somatic embryos and the transformed somatic embryos.
Exposure to 100–1000 mg l−1 of ampicillin, carbenicillin or cefotaxime respectively for up to 60 days did not eliminate the A. tumefaciens while timentin at 500–1000 mg l−1 eradicated it from somatic embryos. One-hour acidified medium treatments and the addition of 100 mg l−1 kanamycin to 500 mg l−1 ampicillin, carbenicillin, cefotaxime or timentin were of little help in eliminating the Agrobacterium. All four antibiotics reduced somatic embryo production, carbenicillin minimally and cefotaxime maximally, especially at
higher concentrations, in comparison with antibiotic-free medium. Putative transformed embryos were selected for continued
proliferation on a 100 mg l−1 kanamycin-containing medium. Histochemical assessments indicated that more gus-positive somatic embryos, particularly fully gus-positive embryos, regenerated from timentin-containing medium than from other antibiotic-containing media under equivalent
conditions. Transformed embryos have been grown and converted into plants and gus activity was observed in whole plants.
Received: 13 July 1999 / Revision received: 2 December 1999 / Accepted: 6 December 1999 |
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Keywords: | Agrobacterium-mediated transformation gus gene expression Juglans regia Timentin Ticarcillin/potassium clavulanate |
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