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Transformation of Papaver somniferum cell suspension cultures with sam1 from A. thaliana results in cell lines of different S-adenosyl-L-methionine synthetase activity

Authors

  • Muriel Belny,

    1. Lab de Biotechnologie et Physiologie Végétales. Fac. des Sciences, Univ. de Picardie Jules Verne, 33 rue Saint Leu, F-80039 Amiens Cedex, France
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  • Didier Hérouart,

    1. Lab. de Biologie Végétale et Microbiologie, URA CNRS 1114, Fac. des Sciences, Parc Valrose, F-06108 Nice Cedex, France
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  • Brigitte Thomasset,

    1. Lab. de Technologie Enzymatique, URA CNRS 1442, UTC, BP 529, F-60205 Compiègne, France
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  • Hélène David,

    1. Lab de Biotechnologie et Physiologie Végétales. Fac. des Sciences, Univ. de Picardie Jules Verne, 33 rue Saint Leu, F-80039 Amiens Cedex, France
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  • Annie Jacquin-Dubreuil,

    1. Lab. de Pharmacognosie et Phytotechnologie, Fac. de Pharmacie, Univ. de Picardie Jules Verne, 1 rue des Louvels, F-80037 Amiens Cedex, France.
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  • Alain David

    Corresponding author
    1. Lab de Biotechnologie et Physiologie Végétales. Fac. des Sciences, Univ. de Picardie Jules Verne, 33 rue Saint Leu, F-80039 Amiens Cedex, France
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(corresponding author, e-mail ah.david@uricardie.fr)

Abstract

We have developed a procedure for opium poppy (Papaver somniferum) transformation using Agrobacterium tumefaciens-mediated gene delivery. Hypocotyl-derived cell suspension cultures of P. somniferum were cocultivated with A. tumefaciens strain GV3101(pMP90) harbouring either the binary vector pTHW136 or the binary vector pO35SSAM. The former contained the uidA reporter gene and the nptII selectable gene, encoding the enzymes β-glucuronidase and neomycin phosphotransferase II, respectively. The latter contained the sam1 gene encoding the enzyme S-adenosyl-L-methionine (SAM) synthetase and the nptII gene. Putatively transformed cell lines were selected on media supplemented with paromomycin. Integration of the foreign genes was confirmed by Southern blot analysis with and without PCR amplification prior to hybridization. SAM synthetase activity was measured in extracts of 5 transformed cell lines. One of them expressed a significant overactivity while two others had a lower activity than the control cell line, leading us to question the possible partial cosuppression of both the resident and the foreign sam genes. To our knowledge, this is the first report of Agrobacterium tumefaciens-mediated transformation of Papaver somniferum cell suspension cultures.

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