Plant Cell, Tissue and Organ Culture

, Volume 74, Issue 1, pp 73–80

Long-term instability of alkaloid production by stably transformed cell lines of Catharanthus roseus

Authors

  • Serap Whitmer
    • Leiden/Amsterdam Center for Drug Research, Division of Pharmacognosy, Gorlaeus LaboratoriesLeiden University
  • Camilo Canel
    • Corning Life Sciences, BF-01
  • Robert van der Heijden
    • Leiden/Amsterdam Center for Drug Research, Division of Pharmacognosy, Gorlaeus LaboratoriesLeiden University
    • Leiden/Amsterdam Center for Drug Research, Division of Pharmacognosy, Gorlaeus LaboratoriesLeiden University
Article

DOI: 10.1023/A:1023368309831

Cite this article as:
Whitmer, S., Canel, C., van der Heijden, R. et al. Plant Cell, Tissue and Organ Culture (2003) 74: 73. doi:10.1023/A:1023368309831

Abstract

The productivity of several transgenic cell lines of Catharanthus roseus was monitored over a period of 30 months. The transgenic cultures were obtained by Agrobacterium-mediated transformation of leaf explants with constructs containing recombinant versions of the endogenous Str and Tdc genes, which, respectively, encode strictosidine synthase (STR) and tryptophan decarboxylase (TDC). The expression of these transgenes and the β-glucuronidase marker gene were also measured periodically, at the enzymatic level, during this time. Cultures were maintained in selective medium containing either hygromycin or kanamycin and showed GUS activity in the presence of X-gluc, indicating that they carried functional transgenes. The activities of STR and TDC varied greatly over time, occasionally falling to levels not significantly different from those of non-transgenic cultures, and showed susceptibility to the composition of the culture medium. Despite maintaining their transgenic character, the cell lines gradually lost the ability to accumulate terpenoid indole alkaloids (TIAs). The diversity of alkaloids produced was also negatively affected by long-term subculture. We conclude that a strategy of indirect selection, such as the use of antibiotic-resistance genes, is insufficient to maintain the concerted expression of TIA-pathway elements necessary for high productivity.

cell culturegenetic engineeringsecondary metabolismstrictosidine synthaseterpenoid indole alkaloidstransgene expressiontryptophan decarboxylase
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© Kluwer Academic Publishers 2003