Abstract
A new method has been developed to introduce foreign DNA into rice cells. Gene delivery occurred when an electrophoretic drive with cycles of intervallic electric field was applied to a mixture containing partially digested small cell groups (SCGs) and plasmid DNAs. Gene transfer efficiency was evaluated by the detection of β-glucuronidase (GUS) activity resulting from expression of a chimaeric plasmid DNA. The optimal combination of treatment conditions (3 V/cm, 30 s pulse and 30 min electrophoretic run) produced a frequency of up to 8.2% of blue cells in transformed microcalluses 40 days after culture of treated SCGs without selection for kanamycin resistance. Southern hybridization showed that the foreign gene had integrated into the chromosomal DNA. These results demonstrate that pulsed electrophoretic drive is applicable to the transfer of foreign genes into plant cells.
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References
Ahokas, H. (1989) Transfer of germinating barley seed electrophoretically with exogenous DNA.Theor. Appl. Genet. 77, 469–72.
Ballas, N., Zakai, N. and Loyter, A. (1987) Transient expression of the plasmid pCaMVCAT in plant protoplasts following transformation with polyethyleneglycol.Exp. Cell Res. 170 228–234.
Batty, N.P. and Evans, J.M. (1992) Biological ballistics — no longer a shot in the dark.Trans. Res. 1, 107–13.
Bevan, M.W. and Chilton, M.D. (1982) T-DNA ofAgrobacterium Ti and Ri plasmids.Annu. Rev. Genet. 16, 357–84.
Christou, P., Ford, T.L. and Kofron, M. (1991) Production of transgenic rice (Oryza sativa L.) plants from agronomically importantindica andjaponica varieties via electric discharge particle acceleration of exogenous DNA into immature zygotic embryos.Bio/Technology 9, 957–62.
Christou, P. (1992) Genetic transformation of crop plants using microprojectile bombardment.Pl. J. 2, 275–81.
Chu, C.C., Wang, C.C., Sun, C.S., Hsu, C., Yin, K.C., Chu, C.Y. and Bi, F.Y. (1975) Establishment of an efficient medium for anther culture of rice through comparative experiments on the nitrogen sources.Scientia Sinica 16, 659–88.
Fraley, R.T., Horsch, R.B., Matzke, A., Chilton, M.D., Chilton, W.S. and Sanders, P.R. (1984)In vitro transformation of petunia cells by an improved method of co-cultivation withA. tumefaciens strains.Pl. Mol. Biol. 3, 371–78.
Frearson, E.M., Power, J.B. and Cocking, E.C. (1973) The isolation culture and regeneration of petunia leaf protoplasts.Dev. Biol. 33, 130–137.
Fromm, M., Taylor, L.P. and Walbot, V. (1985) Expression of gene transfer into monocot and dicot plant cells by electroporation.Proc. Natl Acad. Sci. USA 82, 5824–8.
Hu, C., Chee, P.P., Chesney, R.H., Zhou, J.H., Miller, P.D. and O'Brien, W.T. (1990) Intrinsic GUS-like activities in seed plants.Pl. Cell Rep. 9, 1–5.
Iida, A., Seki, M., Kamada, M., Yamada, Y. and Morikawa, H. (1990) Gene delivery into cultured plant cells by DNA-coated gold particles accelerated by a pneumatic particle gun.Theor. Appl. Genet. 80, 813–6.
Jefferson, R.A. (1987) Assaying chimeric genes in plants: the GUS fusion system.Pl. Mol. Biol. Rep. 5, 387–405.
Junghans, H. and Metzlaff, M. (1990) A simple and rapid method for the preparation of total plant DNA.Bio-Techniques 8, 176.
Klein, T.M., Fromm, M.E., Gradziel, T. and Sanford, J.C. (1988) Factors influencing gene delivery intoZea mays cells by high velocity microprojectiles.Bio/Technology 6, 923–6.
Kyozuka, J., Otoo, E. and Shimamoto, K. (1988) Regeneration from protoplasts ofindica rice: genotypic difference in culture response.Theor. Appl. Genet. 76, 887–90.
Ow, D.W., Wood, K.V., Deluca, M., DeWet, J.R., Heilinski, D.R. and Howell, S.H. (1986) Transient and stable expression of the firefly luciferase gene in plants and transgenic plants.Science 234, 856–9.
Potrykus, I. (1990) Gene transfer to cereals: an assessment.Bio/Technology 8, 535–42.
Rhodes, C.A., Pierce, D.A., Mettler, I.J., Mascarens, D. and Detmer, J.J. (1989). Genetically transformed maize plants from protoplasts.Science 240, 204–7.
Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989)Molecular Cloning: a Laboratory Manual. Second edition. Cold Spring Harbor, New York: Cold Spring Harbor Laboratories.
Sautter, D., Walder, H., Neuhaus-Url, G., Galli, A., Neuhaus, G. and Potrykus, I. (1991) Micro-targetting: high efficiency gene transfer using a novel approach for the acceleration of microprojectiles.Bio/Technology 9, 1080–5.
Shimamoto, K., Terada, R., Izawa, T., Fujimoto, H. (1989) Fertile rice plants regenerated from transformed protoplasts.Nature 338, 274–6.
Southern, E.M. (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis.J. Mol. Biol. 98, 503–17.
Vasil, I.K. (1987) Developing cell and tissue culture for the improvement of cereal and grass crops.J. Pl. Physiol. 128, 193–218.
Yang, J., Ge, K., Lou, M., Wang, Y., Wang, B., Hu, R., Qian, M., Tan, C.C., Lee, N. and Testa, D. (1991) Introduction of foreign genes into small cell groups digested partialenzymatically and regeneration of transgenic plants in rice.Acta Bot. Sinica 3, 825–32.
Zhang, W. and Wu, R. (1988) Efficient regeneration of transgenic plants from rice protoplasts and correctly regulated expression of the foreign gene in the plants.Theor. Appl. Genet. 76, 835–40.
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Yang, J., Ge, K., Wang, Y. et al. Highly efficient transfer and stable integration of foreign DNA into partially digested rice cells using a pulsed electrophoretic drive. Transgenic Research 2, 245–251 (1993). https://doi.org/10.1007/BF01968837
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DOI: https://doi.org/10.1007/BF01968837