Summary
To investigate the possibility, implied by an earlier report, that large amounts of degradable DNA are probably present in the cytoplasm of young cotton embryos, an investigation was undertaken to establish the distribution, amount and metabolic stability of DNA in cotton embryos. Several sensitive cytochemical tests failed to detect any but small amounts of extranuclear DNA. Quantitative determination of the nucleic acid content of embryos during embryogenesis showed that the amounts of DNA and RNA remained fairly constant during embryogenesis, with a ratio of RNA to DNA of about 3.5 to 1. Quantitative autoradiography at both the light and electron microscope levels of sections from embryos pulse-labeled with 3H-thymidine showed that the grain density over the nucleus and cytoplasm did not change during a seven-hour period after labeling, nor did the distribution of label in the cytoplasm. Virtually all incorporation was eliminated by the inclusion of iododeoxy-uridine in the medium. Almost all of the nuclear label and at least 90% of the cytoplasmic label after 3H-thymidine incorporation was eliminated by deoxyribonuclease. It was concluded that there are no unusual features related to DNA distribution or metabolism in cotton embryo; i.e., that only small amounts of DNA are present in the cytoplasm and that all of the DNA is metabolically stable.
Approximately 40% of the cytoplasmic grains after 3H-thymidine labeling were not associated with either plastids or mitochondria (i.e., were more than 0.1 micron distant). No fully satisfactory explanation for such an apparently high figure could be given.
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Branton, D., Ruch, F.: The localization of deoxyribonucleic acid and ultraviolet absorptive granules in Vicia Faba root tips. Exp. Cell Res. 36, 285–296 (1964).
Camargo, E.P., Plant, W.: The radioautographic detection of DNA with tritiated actinomycin D. J. Cell Biol. 35, 713–716 (1967).
Caro, L.G.: High resolution autoradiography II. The problem of resolution. J. Cell Biol. 15, 189–199 (1962).
Caro, L.G., Tubergen, R.P. van, Klab, J.A.: High resolution autoradiography I. Methods. J. Cell Biol. 15, 173–188 (1962).
Ebstein, B.S.: Tritiated actinomycin D as a cytochemical label for small amounts of DNA. J. Cell Biol. 35, 709–712 (1967).
Fisher, D.B., Jensen, W.A., Ashton, M.E.: Histochemical studies of pollen: storage pockets in the endoplasmic reticulum (E. R.). Histochemie 13, 169–182 (1968).
Gahn, P.B., Chaygn, J.: Cytoplasmic deoxyribonucleic acid. Int. Rev. Cytol. 18, 223–247 (1965).
Halperin, W., Wetherell, D.F.: Adventive embryony in tissue cultures of the wild carrot, daucus carota. Amer. J. Bot. 51, 274–283 (1964).
Hatta, Y., Bassil, A., Stern, H.: Nuclear and cytoplasmic DNA from tissues of higher plants. J. Cell Biol. 27, 451–457 (1964).
Iwamura, T.: Nucleic acids in chloroplasts in metabolic DNA. In: Progress in nucleic acid research and molecular biology, ed. by J.N. Davidson, W.E.Colt, vol. 5, p. 133–155. New York-London: Academic Press 1966.
Jensen, W.A.: Botanical histochemistry. San Francisco: W.H. Freeman 1962.
Karnovsky, M.J.: Simple methods for “Staining with Lead” at high pH in electron microscopy. J. of Biophys. and Biochem. 11, 729–732 (1961).
Kasten, F.H.: Schiff-type reagents in cytochemistry I. Theoretical and practical considerations. Histochemie 1, 466–509 (1959).
Keeble, S.A., Joy, R.F.: Fluorescent staining for the differentiation of intracellular ribonucleic acid and deoxyribonucleic acid. Nature (Lond.) 193, 695–696 (1962).
Moses, M.J.: Application of autoradiography to electron microscopy. J. Histochem. Cytochem. 12, 115–130 (1964).
Mourad, E.B.: Evidence for cytoplasmic DNA in root cells of Nicotiana. J. Cell Biol. 24, 267–276 (1965).
Ogur, M., Rasen, G.: The nucleic acids of plant tissues I. The extraction and estimation of desoxypentose nucleic acid and pentose nucleic acid. Arch. Biochem. 25, 262–276 (1950).
Perry, R.R.: Quantitive autoradiography. In: Methods in cell physiology, ed. by D.M. Prescott, vol. 1, p. 305–326. New York: Academic Press 1964.
Raghavan, V., Torrey, J.G.: Growth and morphogenesis of globular and older embryos of Capsella in culture. Amer. J. Bot. 50, 540–551 (1963).
Salpeter, M.M., Beahmann, L.: Autoradiography with the electron microscope. A procedure for improving resolution, sensitivity and contrast. J. Cell Biol. 22, 469–475 (1964).
Sampson, M., Davies, D.D.: Synthesis of a metabolically labile DNA in the maturing root cells of vicia faba. Exp. Cell Res. 43, 669–673 (1966).
Yoo, B., Jensen, W.A.: Changes in nucleic acid content and distribution during cotton embryogenesis. Exp. Cell Res. 42, 447–459 (1966).
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This work was supported by a Public Health Service fellowship 5-F2-GM-22,031-02 from the National Institute of General Medical Sciences, by NSF grant GB 3460, by NIH grant 5-R01-Ca0356-10 and by Miller Institute for Basic Science.
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Fisher, D.B., Jensen, W.A. Nuclear and cytoplasmic DNA synthesis in cotton embryos: a correlated light and electron microscope autoradiographic study. Histochemie 32, 1–22 (1972). https://doi.org/10.1007/BF00277467
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DOI: https://doi.org/10.1007/BF00277467