Summary
Through the use of electroporation and a soybean (Glycine max L.) protoplast system, we generated stably transformed cell lines expressing a number of foreign genes (neomycin phosphotransferase,β-glucuronidase, chloramphenicol acetyl transferase, and phosphinothricin acetyl transferase). Selected and unselected marker genes were cointroduced either linked on a single plasmid or as separate plasmids. Calli expressing multiple genes were recovered, and Cotransformation frequencies were established for both cases. Our results show a 50% cotransformation frequency in the case of linked genes. In situations in which two genes are introduced on independent plasmids, cotransformation frequencies are 18%–27%. Similar rates of cotransformation were observed among various marker pairs.
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References
Bagdasarian M, Lurz R, Ruckert B, Franklin J, Frey J, Bagdasarian M, Timmis K (1981) Specific-purpose plasmid cloning vectors. II. Broad host range, high copy number, RSF 1010-derived vectors, and a host-vector system for gene cloning inPseudomonas. Gene 16:237–247
Barton KA, Whiteley HR, Yang NS (1987)Bacillus thuringiensis delta-endotoxin expressed in transgenicNicotiana tabacum provides resistance to lepidopteran insects. Plant Physiol 85:1103–1109
Bolivar F, Rodriguez RL, Greene PJ, Betlach MC, Heyneker HL, Boyer HW, Crosa JH, Falkow S (1977) Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene 2:95–113
Christou P, Murphy JE, Swain WF (1987) Stable transformation of soybean by electroporation and root formation from transformed callus. Proc Natl Acad Sci USA 84:3962–3966
Christou P, McCabe DE, Swain WF (1988) Stable transformation of soybean callus by DNA-coated gold particles. Plant Physiol 87:671–674
Christou P, Swain WF, Yang NS, McCabe DE (1989) Inheritance and expression of foreign genes in transgenic soybean plants. Proc Natl Acad Sci USA 86:7500–7504
Damm B, Schmidt R, Willmitzer I. (1989) Efficient transformation ofArabidopsis thaliana using direct gene transfer to protoplasts. Mol Gen Genet 217:6–12
De Block M, Botterman J, Vandewiele M, Dockx J, Thoen C, Gossele V, Rao Movva N, Van Montagu M, Leemans J (1987) Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J 6:2513–2518
Gardner R, Howarth A, Hahn P, Brown-Luedi M, Shepherd R, Messing J (1981) The complete nucleotide sequence of an infectious clone of cauliflower mosaic virus by M13mp7 shotgun sequencing. Nucleic Acids Res 9:2871–2888
Gorman C, Moffat I., Howard B (1982) Recombinant genomes which express chloramphenicol acetyl transferase in mammalian cells. Mol Cell Biol 2:1044–1051
Herrera-Estrella I., Depicker A., Van Montagu M, Schell J (1983) Expression of a chimeric gene transferred into plant cells using a Ti-plasmid-derived vector. Nature 303:209–213
Jefferson RA (1987) Assaying chimeric genes in plants. The GUS gene fusion system. Plant Mol Biol Rep 5:387–405
Jefferson RA, Burges SM, Hirsh D (1986)β-Glucuronidase fromE. coli as a gene-fusion marker. Proc Natl Acad Sci USA 83:8447–8451
Jongsma M, Koorneef M, Zabel P, Hille J (1987) Tomato protoplast DNA transformation: Physical linkage and recombination of exogenous DNA sequences. Plant Mol Biol 8:384–394
Krens FA, Mans RMW, Slogteren TMS van, Hoge JHC, Wullems CJ, Schilperoort RA (1985) Structure and expression of DNA transferred to tobacco via transformation of protoplasts with Ti-plasmid DNA: Co-transfer of T-DNA and non-T-DNA sequences. Plant Mol Biol 5:223–224
McCabe DE, Swain WF, Martinell BJ, Christou P (1988) Stable transformation of soybean (Glycine max) by particle acceleration. Bio Technol 6:923–926
Paszkowski J, Schillito RD, Saul M, Mandak V, Hohn T, Hohn B, Potrykus I (1984) Direct gene transfer to plants. EMBO J 3:2717–2722
Peerbolte R, Krens FA, Mans RMW, Floor M, Hoge JHC, Wullems GJ, Schilperoort RA (1985) Transformation of plant protoplasts with DNA: Cotransformation of non-selected calf thymus DNA sequences. Plant Mol Biol 5:235–246
Platt SG, Yang NS (1987) Dot assay for neomycin phosphotransferase activity in crude cell extracts. Ann Biochem 162:529–535
Reiss B, Sprengel R, Will H, Schaller H (1984) A new sensitive method for qualitative and quantitive assay of neomycin phosphotransferase in crude extracts. Gene 30:211–218
Riggs CD, Bates GW (1986) Stable transformation of tobacco by electroporation: Evidence for plasmid concatenation. Proc Natl Aad Sci USA 83:5602–5606
Schocher RJ, Shillito RD, Saul MW, Pasjkowski J, Potrykus I (1986) Cotransformation of unlinked foreign genes into plants by direct gene transfer. Bio/Technol 4:1093–1096
Tagu D, Bergounioux C, Cretin C, Perennes C, Gadal P (1988) Direct gene transfer inPetunia hybrida electroporated protoplasts: Evidence for Cotransformation with a phosphoenolpyruvate carboxylase cDNA from sorghum leaf. Protoplasma 146:101–105
Thompson CJ, Movva NR, Tizard R, Crameri R, Davis JE, Lauwereys M, Botterman J (1987) Characterization of the herbicide-resistant gene BAR fromStreptomyces hygroscopicus. EMBO J 6:2519–2523
Uchimiya H, Hirochika H, Hashimoto H, Hara A, Masuda T, Kasumimoto T, Harada H, Ikeda JE, Yoshioka M (1986) Co-expression and inheritance of foreign genes in transformants obtained by direct DNA transformation of tobacco protoplasts. Mol Gen Genet 205:1–8
Umbeck P, Johnson G, Barton K, Swain W (1987) Genetically transformed cotton (Gossypium hirsutum L.) plants. Bio/ Technol 5:263–266
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Communicated by I. Potrykus
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Christou, P., Swain, W.F. Cotransformation frequencies of foreign genes in soybean cell cultures. Theoret. Appl. Genetics 79, 337–341 (1990). https://doi.org/10.1007/BF01186076
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DOI: https://doi.org/10.1007/BF01186076