Summary
Single cells of hybrid tobacco callus, isolated from germinating seeds, were grown individually in microchambers in the absence of any other cells, in fresh, liquid, coconut milk-medium. These produced a conlony of 50–75 cells in 10–15 days. In all instances the plane of the first cell division, and in most cases also that of the second division, were at right angles to the long axis of the cell. There was more variation in the size, shape and pattern of the second and subsequent divisions in single cells isolated from fresh stem pith and seed callus than those from old stem callus. Upon transfer from the microchambers to agar medium the colony of cells gave rise to a huge mass of callus tissue in about 3 months. In no instance did the cell colonies or any stages preceding them resemble or simulate any stages of normal embryogeny of tabacco. Single-cell clones obtained by this method from seed callus or fresh pith callus produced shoots with leaves and roots in synthetic medium with various indoleacetic acid-kinetin combinations. Normal plants established from the above cultures in the greenhouse flowered in due course of time. This method offers the possibility of producing a very large number of clones or identical plants in species where vegetative propagation is not otherwise possible, apart from its use in studies on the genetics and morphogenesis in higher plants.
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References
Ball, E.: An optically efficient method for observing single plant cells from shake cultures during growth and division. Nature (Lond.) 197, 103–104 (1963).
Bergmann, L.: Growth and division of single cells of higher plants in vitro. J. gen. Physiol. 43, 841–851 (1960).
Blakely, L.M., and F.C. Steward: Growth and organized development of cultured cells. V. The growth of colonies from free cells on nutrient agar. Amer. J. Bot. 51, 780–791 (1964).
Earle, E.D., and J.G. Torrey: Colony formation by isolated Convolvulus cells plated on defined media. Plant Physiol. 40, 520–528 (1965).
Gautheret, R.J.: Sur la possibilité de réaliser la culture indéfinie des tissus de tubercules de carotte. C. R. Acad. Sci. (Paris) 208, 118–120 (1939).
Gibbs, J.L., and D.K. Dougall: Growth of single plant cells. Science 141, 1059 (1963).
Haberlandt, G.: Kulturversuche mit isolierten Pflanzenzellen. S.-B. Akad. Wiss. Wien, math.-nat. Kl. 111, 69–92 (1902).
Halperin, W.: Morphogenetic studies with partially synchronized cultures of carrot embryos. Science 146, 408–410 (1964).
—, and D.F. Wetherell: Adventive embryony in tissue cultures of the wild carrot, Daucus carota. Amer. J. Bot. 51, 274–283 (1964).
——, and D.F. Wetherell Ammonium requirement for embryogenesis in vitro. Nature (Lond.) 205, 519–520 (1965a).
——, and D.F. Wetherell Ontogeny of adventive embryos of wild carrot. Science 147, 756–758 (1965b).
Hildebrandt, A.C.: Tissue and single cell cultures of higher plants as a basic experimental method. In: Modern methods of plant analysis (H.F. Linskens and M.V. Tracey, eds.), vol. 5, p. 383–421. Berlin-Göttingen-Heidelberg: Springer 1962.
Jones, L.E., A.C. Hildebrandt, A.J. Riker, and J.H. Wu: Growth of somatic tobacco cells in microculture. Amer. J. Bot. 47, 468–475 (1960).
Kato, H., and M. Takeuchi: Morphogenesis in vitro starting from single cells of carrot root. Plant and Cell Physiol. 4, 243–245 (1963).
Muir, W.H., A.C. Hildebrandt, and A.J. Riker: Plant tissue cultures produced from single isolated cells. Science 119, 877–878 (1954).
——— The preparation, isolation, and growth in culture of single cells from higher plants. Amer. J. Bot. 45, 589–597 (1958).
Murashige, T., and F. Skoog: A revised medium for a rapid growth and bioassays with tobacco tissue cultures. Physiol. Plantarum (Kbh.) 15, 473–497 (1962).
Reinert, J.: Growth of single cells from higher plants on synthetic media. Nature (Lond.) 200, 90–91 (1963).
Ropp, R.S. De: The growth and behaviour in vitro of isolated plant cells. Proc. roy. Soc. B. 144, 86–93 (1955).
Sievert, R.A., and A.C. Hildebrandt: Variations within single cell clones of tobacco tissue cultures. Amer. J. Bot. 52, 742–750 (1965).
Steward, F.C., L.M. Blakely, A.E. Kent, and M.O. Mapes: Growth and organization in free cell cultures. Brookhaven Symp. Biol. 16, 73–88 (1963).
— M.O. Mapes, A.E. Kent, and R.D. Holsten: Growth and development of cultured plant cells. Science 143, 20–27 (1964).
—— and K. Mears: Growth and organized development of cultured cells. II. Organization in cultures grown from freely suspended cells. Amer. J. Bot. 45, 705–708 (1958).
Torrey, J.G.: Cell division in isolated single plant cells in vitro. Proc. nat. Acad. Sci. (Wash.) 43, 887–891 (1957).
—, and J. Reinert: Suspension cultures of higher plant cells in synthetic media. Plant Physiol. 36, 483–491 (1961).
Vasil, I.K., and A.C. Hildebrandt: Growth and chlorophyll production in plant callus tissues grown in vitro. Planta (Berl.) 68, 69–82 (1966a).
—— Variations of morphogenetic behavior in plant tissue cultures. I. Cichorium endivia Linn. Amer. J. Bot. 53, 860–869 (1966b).
—— Variations of morphogenetic behavior in plant tissue cultures. II. Petroselinum hortense. Amer. J. Bot. 53, 869–874 (1966c).
——, and A.J. Riker: Endive plantlets from freely suspended cells and cell groups grown in vitro. Science 146, 76–77 (1964).
Vasil, Vimla, and A.C. Hildebrandt: Growth and tissue formation from single, isolated tobacco cells in microculture. Science 147, 1454–1455 (1965a).
—— Differentiation of tobacci plants from single, isolated cells in microcultures. Science 150, 889–892 (1965b).
White, P.R.: Potentially unlimited growth of excised plant callus in an artificial nutrient. Amer. J. Bot. 26, 59–64 (1939).
—: Plant tissue culture. Ann. Rev. Biochem. 11, 615–628 (1942).
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Vasil, V., Hildebrandt, A.C. Further studies on the growth and differentiation of single, isolated cells of tobacco in vitro. Planta 75, 139–151 (1967). https://doi.org/10.1007/BF00387129
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DOI: https://doi.org/10.1007/BF00387129