Abstract
Single cells isolated from aggregated Taxus cuspidata cultures via enzymatic digestion were grown in suspension culture. High seeding density (4×105 cells/ml) and the addition of cell-free conditioned medium were essential for growth. Doubling the concentration of the nutrients [ascorbic acid (150 g/l), glutamine (6.25 mm), and citric acid (150 g/l)] had no effect on single cell growth or viability. A specific growth rate of 0.11 days−1 was achieved, which is similar to the observed growth rate of aggregated Taxus suspensions. The biocide, Plant Preservative Mixture, added at 0.2% (v/v) to all single cell cultures to prevent microbial contamination, had no significant effect on growth or viability. Following cell sorting, single cell cultures can be used to establish new cell lines for biotechnology applications or provide cells for further study.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
N Blackhall M Davey J Power (1994) Isolation, culture, and regeneration of protoplasts R Dixon RA Gonzales (Eds) Plant Cell Culture Oxford University Press Oxford 27–39
B Damm R Schmidt L Wilmitzer (1989) ArticleTitleEfficient transformation of Arabidopsis thaliana using direct gene-transfer to protoplasts Mol. Gen. Genet. 217 6–12 Occurrence Handle10.1007/BF00330935 Occurrence Handle2549369
Guri A, Kishor N (1998) Compositions and methods to prevent microbial contamination of plant tissue culture media. United States Patent 5,750,402
SW Hou JF Jia (2004) ArticleTitlePlant regeneration from protoplasts isolated from embryogenic calli of the forage legume Astragalus melilotoides Pall Plant Cell Rep. 22 741–746 Occurrence Handle10.1007/s00299-004-0760-8 Occurrence Handle14762684
R Ketchum D Gibson L Gallo (1995) ArticleTitleMedia optimization for maximum biomass production in cell cultures of pacific yew Plant Cell Tiss. Org. 42 185–193 Occurrence Handle10.1007/BF00034237
P King K Mansfield H Street (1973) ArticleTitleControl of growth and cell division in plant cell suspension cultures Can. J. Bot. 51 1807–1823
D Kubek M Shuler (1978) ArticleTitleOn the generality of methods to obtain single-cell plant suspension cultures Can. J. Bot. 56 2521–2527
W Kurz (1971) ArticleTitleA chemostat for growing higher plant cells in single cell suspension cultures Exp. Cell Res. 64 476–9 Occurrence Handle10.1016/0014-4827(71)90104-2 Occurrence Handle5542654
C Lee M Shuler (2000) ArticleTitleThe effect of inoculum density and conditioned medium on the production of ajmalicine and catharanthine from immobilized Catharanthus roseus cells Biotechnol. Bioeng. 67 61–71 Occurrence Handle10.1002/(SICI)1097-0290(20000105)67:1<61::AID-BIT7>3.0.CO;2-J Occurrence Handle10581436
R Ma YD Guo S Pulli (2003) ArticleTitleSomatic embryogenesis and fertile green plant regeneration from suspension cell-derived protoplasts of rye (Secale cereale L.) Plant Cell Rep. 22 320–327 Occurrence Handle10.1007/s00299-003-0694-6 Occurrence Handle14648108
M Naill S Roberts (2004) ArticleTitlePreparation of isolated cells from Taxus supension cultures for population analysis Biotechnol. Bioeng. 86 817–826 Occurrence Handle10.1002/bit.20083 Occurrence Handle15162458
M Naill S Roberts (2005a) ArticleTitleFlow cytometric analysis of protein content in Taxus protoplasts and single cells as compared to aggregated suspension cultures Plant Cell Rep. 23 528–533 Occurrence Handle10.1007/s00299-004-0875-y
M Naill S Roberts (2005b) ArticleTitleCell cycle analysis of Taxus suspension cultures at the single cell level as an indicator of culture heterogeneity Biotechnol. Bioeng. 90 491–500 Occurrence Handle10.1002/bit.20446
M Naill S Roberts (2005c) ArticleTitleFlow cytometric identification of paclitaxel accumulating subpopulations Biotechnol. Prog. 21 978–983 Occurrence Handle10.1021/bp049544l
L Pestchanker SC Roberts M Shuler (1996) ArticleTitleKinetics of taxol production and nutrient use in suspension cultures of Taxus cuspidata in shake flasks and a Wilson-type bioreactor Enzyme Microb. Technol. 19 256–260 Occurrence Handle10.1016/0141-0229(95)00243-X Occurrence Handle8987485
S Roberts M Naill D Gibson M Shuler (2003) ArticleTitleA simple method for enhancing paclitaxel release from Taxus canadensis cell suspension cultures utilizing cell wall digesting enzymes Plant Cell Rep. 21 1217–1220 Occurrence Handle10.1007/s00299-003-0575-z Occurrence Handle12811504
K Sakamoto K Iida T Koyano Y Asada T Furuya (1994) ArticleTitleMethod for selecting anthocyanin producing cells by a cell sorter Planta Med. 60 253–259 Occurrence Handle8073093
V Srinivasan L Pestchanker S Moser T Hirasuna R Taticek M Shuler (1995) ArticleTitleTaxol production in bioreactors: kinetics of biomass accumulation, nutrient uptake, and taxol production by cell suspension of Taxus baccata Biotechnol. Bioeng. 47 666–676 Occurrence Handle10.1002/bit.260470607
H Street (1977) Single cell clones-derivation and selection H, Street (Eds) Plant Tissue and Cell Culture Blackwell Scientific Oxford 207–222
H Street G Henshaw M Buiatti (1965) ArticleTitleThe culture of isolated plant cells Chem. Ind. 1 27–33
L Wang J Sun M Horvat N Koutalistras B Johnston A Sheil (1996) ArticleTitleEvaluation of MTS, XTT, MTT, and 3HTdR incorporation for assessing hepatocyte density,␣viability, and proliferation Meth. Cell Sci. 18 249–255 Occurrence Handle10.1007/BF00132890
W Yanpaisan N King P Doran (1999) ArticleTitleFlow cytometry of plant cells with applications in large-scale bioprocessing Biotechnol. Adv. 17 3–27 Occurrence Handle10.1016/S0734-9750(98)00014-7 Occurrence Handle14538141
XY Zhang G Le Pennec R Steffen WEG Muller W Zhang (2004) ArticleTitleApplication of a MTT assay for screening nutritional factors in growth media of primary sponge cell culture Biotechnol. Prog. 20 151–155 Occurrence Handle10.1021/bp0341601 Occurrence Handle14763838
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Naill, M.C., Roberts, S.C. Culture of Isolated Single Cells from Taxus Suspensions for the Propagation of Superior Cell Populations. Biotechnol Lett 27, 1725–1730 (2005). https://doi.org/10.1007/s10529-005-2738-1
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s10529-005-2738-1