Summary
Conditions faborable for the uptake of artificial lipid vesicles (liposomes) encapsulating plasmid DNA byDaucus carota protoplasts were investigated. Incubation period necessary for the maximum uptake of liposome-DNA was in the neighborhood of 10 minutes under the circumstances where liposomes (approximately 1.28 μmoles lecithin/ml) were mixed with 5 × 106 protoplasts/ml. Results obtained by Southern hybridization indicated that the recombinant DNA vector, pBR325 was found in protoplasts after 20 hours incubation in the open circular, linear and some complexed forms. There were some variations in DNA-uptake among protoplasts from different plant species. The gradual disappearance of plasmid molecules was confirmed after 1 week culture, suggesting instability of pBR325 in prolonged cell culture.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Lurquin, P. F., 1979: Entrapment of plasmid DNA by liposomes and their interaction with plant protoplasts. Nucleic Acids Res.6, 3773–3784.
—, 1981: Binding of plasmid loaded liposomes to plant protoplasts: Validity of biochemical methods to evaluate the transfer of exogenous DNA. Plant Sci. Lett.21, 31–40.
Matthews, B. F., Cress, D. E., 1981: Liposome-mediated delivery of DNA to carrot protoplasts. Planta153, 90–94.
Murashige, T., Skoog, F., 1962: A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant.15, 473–479.
Rigby, P. W. J., Dieckmann, M., Rhodes, C., Berg, P., 1977: Labelling deoxyribonucleic acid to high specific activityin vitro by nick translation with DNA and DNA polymerase 1. J. molec. Biol.113, 237–251.
Rollo, F., Galli, M. G., Parisi, B., 1981: Liposome-mediated transfer of DNA to carrot protoplasts: A biochemical and autoradiographic analysis. Plant Sci. Lett.20, 347–354.
Southern, E., 1975: Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. molec. Biol.98, 503–517.
Uchimiya, H., 1981: Parameters influencing the liposome-mediated insertion of fluorescein diacetate into plant protoplasts. Plant Physiol.67, 629–632.
—,Harada, H., 1981: Transfer of liposome-sequestering plasmid DNA intoDaucus carota protoplasts. Plant Physiol.68, 1027–1030.
—,Murashige, T., 1977: Quantitative analysis of the fate of exogenous DNA inNicotiana protoplasts. Plant Physiol.59, 301–308.
Wahl, G. M., Stern, M., Stark, G. R., 1979: Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc. nat. Acad. Sci. U.S.A.76, 3683–3687.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Ohgawara, T., Uchimiya, H. & Harada, H. Uptake of liposome-encapsulating plasmid DNA by plant protoplasts and molecular fate of foreign DNA. Protoplasma 116, 145–148 (1983). https://doi.org/10.1007/BF01279831
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF01279831