Summary
A stable intergeneric transfer of nuclear genes from Physalis minima into the genome of Datura innoxia has been achieved through asymmetric protoplast fusion. No hybrid plants could be obtained from these species either by traditional methods of sexual breeding or by somatic hybridization via fusion of protoplasts containing complete nuclear genomes. The incompatibility barriers were bypassed by the fusion of highly X-irradiated (LD100) wild-type Physalis with nuclear albino mutant Datura protoplasts. In this intergeneric reconstruction, 1.15% of the total heterokaryons restored the chlorophyll synthesis in their regenerants. Two representative transformed lines, TRL-A and TRL-D, were further characterized, showing 0.43–0.78 pg. additional nuclear DNA (4.45–8.07% nuclear DNA of P. minima). Since chromosomes of the species of Datura and Physalis were distinguishable, the mitotic complement of the transformed regenerants showed only 3 and 1 chromosomes of the donor in tetraploid (2n=48) and octoploid (2n=96) genomes of the recipient, respectively. The introduction and expression of limited genes of Physalis in Datura have also been confirmed by the allelic expression of various isoenzymes. Such stable gene transfer via asymmetric fusion of protoplasts has been discussed in relation to its application in the genetic manipulation of plants.
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References
Binding H (1974) Regeneration von haploiden und diploiden Pflanzen aus Protoplasten von Petunia hybrida L. Z Pflanzenphysiol 74:327–356
Binding H, Nehls R (1978) Somatic cell hybridization of Vicia faba+Petunia hybrida. Mol Gen Genet 164:137–143
Binding H, Nehls R (1983) Techniques of somatic cell hybridization by fusion of protoplasts. In: Shay JW (ed) Techniques in somatic cell genetics. Plenum Press, New York, p 471–492
Brewbaker JL, Upadhya MD, Mäkinen Y, Macdonald T (1968) Isoenzyme polymorphism in flowering plants III. Gel electrophoretic methods and applications. Physiol Plant 21:930–940
Davis BJ (1964) Disc electrophoresis II. Method and application to human serum proteins. Ann NY Acad Sci 121:404–427
Dudits D, Hadlaczky G, Bajszár GY, Koncz C, Lázár G, Horváth G (1979) Plant regeneration from intergeneric cell hybrids. Plant Sci Lett 15:101–112
Dudits D, Fejér O, Hadlaczky G, Koncz C, Lázár G, Horváth G (1980) Intergeneric gene transfer mediated by plant protoplast fusion. Mol Gen Genet 179:283–288
Galun E (1982) Somatic cell fusion for inducing cytoplasmic exchange: A new biological system for cytoplasmic genetics in higher plants. In: Vasil IK, Scowcroft WR, Frey KJ (eds) Plant improvement and somatic cell genetics. Academic Press, New York, p 205–219
Gupta PP, Gupta M, Schieder O (1982) Correction of nitrate reductase defect in auxotrophic plant cells through protoplastmediated intergeneric gene transfers. Mol Gen Genet 188:378–383
Kao KN (1977) Chromosomal behaviour in somatic hybrids of soybean-Nicotiana glauca. Mol Gen Genet 150:225–230
Kao KN, Michayluk MR (1974) A method for high frequency integeneric fusion of plant protoplasts. Planta 115:355–367
Keller WA, Setterfield G, Douglas G, Gleddie S, Nakamura C (1982) Production, characterization, and utilization of somatic hybrids of higher plants. In: Tomes DT, Ellis BE, Harney PM, Kasha KJ, Peterson RL (eds) Application of plant cell and tissue culture to agriculture and industry. Guelph University Press, Guelph, p 81–114
Keller WA, Melchers G (1973) The effect of high pH and calcium on tobacco leaf protoplast fusion. Z Naturforsch 28:737–741
Krumbiegel G, Schieder O (1981) Comparison of somatic and sexual incompatibility between Datura innoxia and Atropa belladona. Planta 153:466–470
Laycock MY, Thurman DA, Bouler D (1965) An improved method for the detection of dehydrogenases using tetrazolium salts. Clin Chim Acta 11:98–100
Le John HB, Suzuki I, Wright JA (1968) Glutamate dehydrogenases of Thiobacillus novellus. Kinetic properties and a possible control mechanism. J Biol Chem 243:118–128
Lin MS, Comings DE, Alfi OS (1977) Optical studies of the interaction of 4′-6-diamidino-2-phenylindole with DNA and metaphase chromosomes. Chromosoma (Berl) 60:15–25
Mackinney G (1941) Absorption of light by chlorophyll solutions. J Biol Chem 140:315–322
Melchers G, Sacristán MD, Holder AA (1978) Somatic hybrid plants of potato and tomato regenerated from fused protoplasts. Carlsberg Res Commun 43:203–218
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiol Plantarum 15:473–597
Schieder O (1976) Isolation of mutants with altered pigments after irradiating haploid protoplasts from Datura innoxia Mill with X-rays. Mol Gen Genet 149:251–254
Schieder O (1977) Hybridization experiments with protoplasts from chlorophyll-deficient mutants of some Solanaceous species. Planta 137:253–257
Schieder O (1978) Genetic evidence for the hybrid nature of somatic hybrids from Datura innoxia Mill. Planta 141:333–334
Schieder O, Gupta PP, Krumbiegel G, Hein T (1983) Protoplast fusion and transformation. In: Better crops for food. Pitman Books, London (Ciba Foundation Symp 97), p 213–227
Sehiller B, Herrmann RG, Melchers G (1982) Restriction endonuclease analysis of plastid DNA from tomato, potato and some of their somatic hybrids. Mol Gen Genet 186:453–459
Zenkteler M, Melchers G (1978) In vitro hybridization by sexual methods and by fusion of somatic protoplasts. Theor Appl Genet 52:81–90
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Gupta, P.P., Schieder, O. & Gupta, M. Intergeneric nuclear gene transfer between somatically and sexually incompatible plants through asymmetric protoplast fusion. Mol Gen Genet 197, 30–35 (1984). https://doi.org/10.1007/BF00327919
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DOI: https://doi.org/10.1007/BF00327919