Summary
The binding of the14C-labelledSalmonella typhimurium DNA or3H-labelled soybean SB-1 DNA to cultured soybean cells (Glycine max L. Merr.) (SB-1) could be increased at least 100-fold by choosing the proper incubation conditions. The uptake of DNA by cells could completely be inhibited by the addition of an excess of unlabelled thymidine, indicating that the observed uptake of DNA by cells most probably is simply uptake of DNA degradation products. Autoradiograms, prepared from SB-1 protoplasts that were previously incubated with DNA, showed that the DNA was not associated with the protoplasts, but only with aggregates of cell wall material contaminating the protoplast preparation. When protoplasts and DNA were incubated in the presence of DEAE-dextran, the amount of DNAse resistant radioactivity increased 40 times. Again, the autoradiograms showed that most if not all DNAse-resistant material was associated with cell wall materials. Our observation that it is cell wall contaminants in protoplast preparations which account for most of the DNA binding demonstrates the need for caution in interpreting experiments on the binding and uptake of DNA by plant protoplasts.
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Bendich, A. J., Filner, P., 1971: Uptake of exogenous DNA by pea seedlings and tobacco cells. Mutation Res.13, 199–214.
Chu, K., Lark, K. G., 1976: Cell cycle parameters of soybean (Glycine max L.) cells growing in suspension cultures: suitability of the system for genetic studies. Planta132, 259–268.
Gamborg, O. L., 1970: The effects of amino acids and ammonium on the growth of plant cells in suspension cultures. Plant Physiol.45, 372–375.
Hess, D., 1969: Versuche zur Transformation an höheren Pflanzen: Induktion und konstante Weitergabe der Anthocyansynthese beiPetunia hybrida. Z. Pflanzenphysiol.60, 345–358.
Heyn, R. F., Schilperoort, R. A., 1973: The use of protoplasts to follow the fate ofAgrobacterium tumefaciens DNA on incubation with tobacco cells. Coll. intern. C.N.R.S.212, 385–395.
Holl, F. B., Gamborg, O. L., Ohyama, K., Pelcher, L. E., 1974: Genetic transformation in plants, in: Tissue culture and plant science (Street, H. E., ed.), pp. 301–327. New York: Academic Press Inc.
Ledoux, L., Huart, R., 1969: Fate of exogenous bacterial deoxyribonucleic acids in barley seedlings. J. molec. Biol.43, 243–262.
— —, 1974: DNA-mediated genetic correction of thiaminelessArabidopsis thaliana. Nature249, 17–21.
Lurquin, P. F., Hotta, Y., 1975: Reutilization of bacterial DNA byArabidopsis thaliana cells in tissue cultures. Plant Science Lett.5, 103–112.
—,Kado, C. I., 1977:Escherichia coli plasmid p BR 313 insertion into plant protoplasts and into their nuclei. Molec. gen. Genet.154, 113–121.
Merril, C. R., Stanbro, H., 1974: Intercellular gene transfer. Z. Pflanzenphysiol.73, 371–388.
Nagata, T., Takebe, I., 1970: Cell wall regeneration and cell division in isolated tobacco mesophyll protoplasts. Planta92, 301–308.
Ohyama, K., Gamborg, O. L., Miller, R. A., 1972a: Uptake of exogenous DNA by plant protoplasts. Canad. J. Bot.50, 2077–2088.
— — —, 1972b: Isolation and properties of DNA from protoplasts of cell suspension cultures ofAmmi visnaga and carrot (Daucus carota). Plant Physiol.50, 319–321.
Suzuki, M., Takebe, L., 1976: Uptake of single-stranded bacteriophage DNA by isolated tobacco protoplasts. Z. Pflanzenphys.78, 421–433.
Tomasz, A., 1969: Some aspects of the competent state in genetic transformation. Ann. Rev. Genet.3, 217–232.
Uchimiya, H., Murashige, T., 1976: Quantitative analysis of the fate of exogenous DNA inNicotiana protoplasts. Plant Physiol.59, 301–308.
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NRCC No. 16353.
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Kool, A.J., Pelcher, L.E. Conditions that affect binding of DNA by cultured plant cells and protoplasts: An autoradiographic analysis. Protoplasma 97, 71–84 (1978). https://doi.org/10.1007/BF01276391
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DOI: https://doi.org/10.1007/BF01276391