Selection of high-yielding cell line and establishment of high Taxol-producing cell suspension culture in European yew tree (Taxus baccata L.)

Document Type : Scientific article

Authors

1 PhD Student of Molecular Genetics and Genetic Engineering, Department of Genetics and Plant Production, Faculty of Agriculture, Urmia University, Urmia, I. R. Iran

2 Associate Professor, Department of Genetics and Plant Production, Faculty of Agriculture, Urmia University, Urmia, I. R. Iran

Abstract

Background and Objective: European yew (Taxus baccata L.) is a slow-growing, evergreen conifer native to northern Iran with high medicinal value due to its production of paclitaxel (Taxol), a vital anticancer drug. However, the extremely low concentration of paclitaxel in yew tissues (0.001–0.02% of dry weight) poses a major barrier to sustainable exploitation. Producing just one kilogram of paclitaxel requires about 10,000 kg of bark or the felling of 3,000 trees, while treating a single cancer patient needs approximately 3 g of paclitaxel—equivalent to eight 60-year-old trees. Such demand threatens the survival of natural yew forests. Consequently, biotechnologists seek alternative strategies, such as cell and tissue culture, for large-scale production of secondary metabolites without environmental damage. Cell suspension culture is considered an effective approach for producing medicinal metabolites under controlled laboratory conditions. Establishing high-yield, selected cell lines can significantly enhance biomass and paclitaxel production. This study aimed to develop efficient paclitaxel-producing suspension cultures of T. baccata by selecting high-yield cell lines through small cell aggregate cloning.
Material and Methods: Stem explants from one-year-old yew seedlings were cultured on B5 medium supplemented with various auxins (2,4-D, PIC, NAA) and cytokinins (BAP, KIN) to induce callus formation. Successful induction and optimal phytohormone combinations were determined based on induction time, callus morphology (texture and color), and callogenic response. Selected calli were used to establish primary cell suspension cultures. High-yield cell lines were isolated by cloning small cell aggregates. Resulting suspension cultures from selected lines were analyzed for growth kinetics, paclitaxel accumulation, growth stability, and metabolite production. All experiments were conducted in a completely randomized design with three replicates, repeated twice, and analyzed using SAS 9.4.
Results: Phytohormones significantly influenced callogenesis, with PIC and KIN producing friable, fast-growing callus. Four single-cell-derived clones with superior growth performance were selected from T. baccata suspension cultures. Growth profiling revealed an exponential phase up to day 21, followed by a stationary phase until day 28. Paclitaxel production peaked at the late exponential phase (day 21), coinciding with maximum cell growth. The best-performing line produced the highest paclitaxel yield (26.07 mg/L) and biomass (33.68 g DW/L), about 15-fold and 2.05-fold higher, respectively, than the low-yield line. Cell population dynamics indicated that the superior line predominantly consisted of spherical cell aggregates with rapid division rates and high viability. Stability analysis showed significant differences among lines across subcultures, with only line TbCL-J1 maintaining high productivity and stability in biomass and paclitaxel yield after five successive subcultures.
Conclusion: This study successfully established a high-yield cell line and developed efficient T. baccata cell suspension cultures for paclitaxel production. Paclitaxel yield in the selected line (0.77% of dry biomass) was dramatically higher than that reported for natural yew trees (0.001–0.02%). The cell line selection strategy not only boosted paclitaxel yield but also enhanced biomass accumulation. The small-aggregate cloning method offers potential for application in other plant culture systems. Moreover, the homogeneous cultures derived from single-cell selection in T. baccata proliferate more rapidly than wild trees, making them suitable for scale-up, metabolic engineering, and industrial bioreactor applications—without destructive harvesting of Iran’s native forests.

Keywords

References

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Forest Research and Development
Volume 11, Issue 2
September 2025
Pages 183-204

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