Summary
Protoplasts from callus tissue of Arabidopsis thaliana and from leaves of Brassica campestris were fused using the PEG-high pH-high Ca++-technique. Single dividing fusion products were isolated mechanically and cultured further in 1 μl microdroplets. 31 cell lines each originating from a single fusion event have been obtained. Cytological analysis of 6 cell lines was conducted 4–7 months after isolation. All metaphases examined contained chromosomes of both species. Both Arabidopsis and Brassica specific chromosomes were still retained in cells of 7 month old cultures. Reconstituted di- and multiconstrictional chromosomes were also observed. Biochemical analysis of 12 cell lines was performed in 6–7-month old cultures. Electrophoresis on PAG and specific staining for esterase, lactate dehydrogenase, and peroxidase activities revealed isozymes of both parents to be present in all cell lines. The data obtained are interpreted as evidence for retention and functional activity in hybrid cells of specific chromosomes from each species for at least 7 months of culture.
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Ambros, P., Schweizer, D.: The Giemsa C-banded karyotype of Arabidopsis thaliana (L.) Heynh. Arabidopsis Inform. Serv. 13, 167–171 (1976)
Bobrow, M., Cross, J.: Differential staining of human and mouse chromosomes in interspecific cell hybrids. Nature (Lond.) 251, 77–79 (1974)
Constabel, F.: Isolation and culture of plant protoplasts. In: Plant tissue culture methods (O.L. Gamborg and L.R. Wetter, eds.) pp. 11–16. Saskatoon: NRCC 1975
Davis, B.J.: Disc electrophoresis. II. Method and application to human serum proteins. Ann. N.Y. Acad. Sci. 121, 404–427 (1964)
Galun, E., Raveh, D.: In vitro culture of tobacco protoplasts: survival of diploid and haploid protoplasts exposed to X-ray radiation at different times after isolation. Rad. Botany 15, 79–82 (1975)
Gamborg, O.L., Miller, R.A., Ojima, K.: Nutrient requirements of suspension of soybean root cells. Exp. Cell Res. 50, 151–158 (1968)
Gerstel, D.U., Burns, J.A.: Chromosomes of unusual length in hybrids between two species of Nicotiana. In: Chromosomes today (C.D. Darlington and K.R. Lewis, eds.) pp. 41–56. Edinbourgh: Oliver & Boyd 1966
Gleba, Yu.: Microdroplet culture. Tobacco plants from single mesophyll protoplasts. Naturwissenschaften 65, 158–159 (1978)
Gleba, Yu., Sidorov, V.A., Artemenko, V.S.: Cultivation and fusion of protoplasts isolated from callus of Arabidopsis thaliana. Soviet Plant Physiol. 25, 161–165 (1978)
Kao, K.N.: A chromosomal staining method for cultured cells. In: Plant tissue culture methods (O.L. Gamborg and L.R. Wetter, eds.), pp. 63–64. Saskatoon: NRCC 1975
Kao, K.N.: Chromosomal behaviour in somatic hybrids of soybean-Nicotiana glauca. Molec. gen. Genet. 150, 225–230 (1977)
Kao, K.N., Michayluk, M.R.: A method for high-frequency intergeneric fusion of plant protoplasts. Planta (Berl.) 115, 355–367 (1974)
Kao, K.N., Michayluk, M.R.: Nutritional requirements for growth of Vicia hajastana cells and protoplasts at a very low population density in liquid media. Planta (Berl.) 126, 105–109 (1975)
Kasha, K.J., Kao, K.N.: High frequency of haploid production in barley (Hordeum vulgare L.) Nature (Lond.) 225, 874–875 (1970)
Keller, W.A., Melchers, G.: The effect of high pH and calcium on tobacco leaf protoplasts fusion. Z. Naturforsch. 28c, 737–741 (1973)
Maurer, H.R.: Disk-Elektrophorese. Berlin: Walter de Gruyter & Co. 1968
Murashige, T., Skoog, F.: A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol. Plant. 15, 473–497 (1962)
Nagata, T., Takebe, I.: Plating of isolated tobacco mesophyll protoplasts on agar medium. Planta (Berl.) 99, 12–20 (1971)
Schenck, H., Hoffmann, F.: Callus and root regeneration from mesophyll protoplasts of basic Brassica species: B. campestris, B. oleracea, B. nigra. Z. Pflanzenzüchtg., 1978 (in press)
Schieder, O.: Somatic hybrids of Datura innoxia Mill. + Datura discolor Berak. and of Datura stramonium L. var. torula I. Selection and Characterization. Molec. gen. Genet. 162, 113–119 (1978)
Wetter, L.R.: Isoenzyme patterns in soybean-Nicotiana somatic hybrid cell lines. Molec. gen. Genet. 150, 231–235 (1977)
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Communicated by G. Melchers
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Gleba, Y.Y., Hoffmann, F. Hybrid cell lines Arabidopsis thaliana + Brassica campestris: No evidence for specific chromosome elimination. Molec. Gen. Genet. 165, 257–264 (1978). https://doi.org/10.1007/BF00332524
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DOI: https://doi.org/10.1007/BF00332524