Abstract
Image analysis has been used to assess the growth of cell suspension-derived protoplasts of Petunia hybrida cv. Comanche at an interface between aqueous culture medium (KM8P), supplemented with 0.01% (w/v) Pluronic F-68, and oxygenated (10 mbar; 10 min) perfluorodecalin. Protoplasts synthesised a new cell wall and entered normal mitotic division which was sustainable to the cell colony/callus stage. This process was accentuated by the collective and additive effects of oxygen, perfluorodecalin and surfactant media supplements. The mean area (mm3) of protoplast-derived cell colonies after 68 days of growth was increased 35 fold over control (media alone) in the presence of these combined treatments. The new cultural regime, leading to improved cell throughput from protoplasts, is discussed primarily in relation to the role of perfluorodecalin as a gas carrier and possible effects of Pluronic F-68 in stimulating cellular uptake of nutrients and/or growth regulators. Image analysis provides a novel and accurate approach to quantifying cell growth responses.
Similar content being viewed by others
Abbreviations
- dpi:
-
dots per inch
- FPE:
-
final plating efficiency
- IPE:
-
initial plating efficiency
- KM:
-
Kao & Michayluk (1975)
- PFC:
-
Perfluorocarbon
- UM:
-
Uchimiya & Murashige (1974)
References
Anthony P, Davey MR, Power JB, Washington C & Lowe KC (1994) Synergistic enhancement of protoplast growth by oxygenated perfluorocarbon and Pluronic F-68. Plant Cell Rep. 13: 251–255
Gilmour DM, Golds TJ & Davey MR (1989) Medicago protoplasts: fusion, culture and plant regeneration. In: Bajaj YPS (Ed) Biotechnology in Agriculture and Forestry. Plant Protoplasts and Genetic Engineering I, Vol 8 (pp 370–388). Springer-Verlag, Heidelberg
Harrell RC, Bieniek M & Cantliffe DJ (1992) Noninvasive evaluation of somatic embryogenesis. Biotechnol. Bioeng. 39: 378–383
Ju L-K & Armiger WB (1992) Use of perfluorocarbon emulsions in cell culture. Bio Techniques 12: 258–263
Ju L-K, Lee JF & Armiger WB (1991) Enhancing oxygen transfer in bioreactors by perfluorocarbon, emulsions. Biotechnol. Prog. 7: 323–329
Junker BH, Hatton TA & Wang DIC (1990) Oxygen transfer enhancement in aqueous/perfluorocarbon fermentation systems. I. Experimental observations. Biotechnol, Bioeng. 35: 578–585
Kao KN & Michayluk MR (1975) Nutritional requirements for growth of Vicia hajastana cells and protoplasts at a very low population density in liquid media. Planta 126: 105–110
King AT, Mulligan BJ & Lowe KC (1989) Perfluorochemicals and cell culture. Bio/Technol. 7: 1037–1042
Latif M, Mumtaz N, Davey MR & Power JB (1993) Plant regeneration from protoplasts isolated from cell suspension cultures of the wild tomato, Lycopersicon chilense Dun. Plant Cell Tiss. Org. Cult. 32: 311–317
Lowe KC, Davey MR, Power JB & Mulligan BJ (1993) Surfactant supplements in plant culture systems. Agro-food-Industry Hi-Tech 4: 9–13
Pons M-N, Wagner A, Vivier H & Marc A (1992) Application of quantitative image analysis to a mammalian cell line grown on microcaniers. Biotechnol. Bioeng. 40: 187–193
Power JB, Davey MR, McLellan M & Wilson D (1990) Isolation, culture and fusion of protoplasts-2. Fusion of protoplasts. Biotechnol. Educat. 1: 115–124
Reichl U, King R & Gilles ED (1992) Characterisation of pellet morphology during submerged growth of Streptomyces tendae by image analysis. Biotechnol, Bioeng. 39: 164–170
Rols JL & Goma G (1989) Enhancement of oxygen transfer rates in fermentation using oxygen-vectors. Biotech. Adv 7: 1–14
Serra J (1972) Stereology and structuring elements. J. Micros. 95: 93–103
Snedecor GW & Cochran WG (1989) Statistical Methods, 8th Edition. Ames, Iowa State University Press
Uchimiya H & Murashige T (1974) Evaluation of parameters in the isolation of viable protoplasts from cultured tobacco cells. Plant Physiol. 54: 936–944
Widholm J (1972) The use of FDA and phenosafranine for determining viability of cultured plant cells. Stain, Technol, 47: 186–194
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Anthony, P., Davey, M.R., Power, J.B. et al. Image analysis assessments of perfluorocarbon- and surfactant-enhanced protoplast division. Plant Cell Tiss Organ Cult 38, 39–43 (1994). https://doi.org/10.1007/BF00034441
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00034441