Skip to main content
SpringerLink
Log in

Phleomycin resistance as a dominant selectable marker for plant cell transformation

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

Tobacco cells are sensitive to bleomycin and phleomycin. The Tn5 and the Streptoalloteichus hindustanus (Sh) bleomycin resistance (‘Ble’) genes conferring resistance to these antibiotics have each been inserted into two plant expression vectors. They are flanked by the nopaline synthase (nos) or the cauliflower mosaic virus (CaMV) 35S promoters on one side, and by the nos polyadenylation signal on the other. These four chimaeric genes were introduced into the binary transformation vector pGA 492, which were thereafter mobilized into Agrobacterium tumefaciens strain LBA 4404. The resulting strains were used to transform Nicotiana tabacum cv. Xanthi nc using the leaf disc transformation procedure. In all cases, phleomycin- and bleomycin-resistant tobacco plants were regenerated from transformed cells under selective conditions; however the highest frequency of rooted plants was obtained when transformation was carried out with the ‘Sh Ble’ gene under the control of the 35S promoter. Phleomycin resistance was stably transmitted to sexual offspring as a dominant nuclear trait as confirmed by Southern blotting.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. An G: Development of plant promoter expression vectors and their use for analysis of differential activity of nopaline synthase promoter in transformed tobacco cells. Plant Physiol 81: 86–91 (1986).

    Google Scholar 

  2. An G, Watson BD, Stachel S, Gordon MP, Nester EW: New Cloning vehicles for transformation of higher plants. EMBO J 4: 277–284 (1985).

    Google Scholar 

  3. Bevan M: Binary Agrobacterium vectors for plant transformation. Nucleic Acids Res 12: 8711–8721 (1984).

    Google Scholar 

  4. Bevan M, Barnes WM, Chilton MD: Structure and transcription of the nopaline synthase gene region of T-DNA. Nucleic Acids Res 11: 369–385 (1983).

    PubMed  Google Scholar 

  5. Bevan MW, Flavell RB, Chilton MD: A chimaeric antibiotic resistance gene as a selectable marker for plant cell transformation. Nature 304: 184–187 (1983).

    Google Scholar 

  6. Birnboim HC, Doly J: A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7: 151–1523 (1979).

    PubMed  Google Scholar 

  7. Boyer HW, Roulland-Dussoix D: A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol 41: 459–472 (1969).

    Article  PubMed  Google Scholar 

  8. DeBlock M, Botterman J, Vandewiele M, Dockx J, Thoen C, Gosselé V, Hovva N, Thompson C, VanMontagu M and Leemans J: Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J 6: 2513–2518 (1987).

    Google Scholar 

  9. Dellaporta SL, Wood J, Hicks JB: A plant DNA minipreparation: version II. Plant Mol Biol Rep 1: 19–21 (1983).

    Google Scholar 

  10. Ditta G, Stanfield S, Corbin D, Helinski DR: Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium melitoti. Proc Natl Acad Sci USA 77: 7347–7351 (1980).

    PubMed  Google Scholar 

  11. Everett NP, Robinson KEP, Mascarenhas D: Genetic engineering of sunflower (Helianthus annuus L.). Biotechnology 5: 120–1204 (1987).

    Article  Google Scholar 

  12. Fraley RT, Rogers SG, Horsch RB, Sanders PR, Flick JS, Adams SP, Bittner ML, Brand LA, Fink CL, Fry JS, Galluppi GR, Goldberg SB, Hoffmann NL, Woo SC: Expression of bacterial genes in plant cells. Proc Natl Acad Sci USA 80: 4803–4807 (1983).

    PubMed  Google Scholar 

  13. Franck A, Guilley H, Jonard G, Richards K, Hirth L: Nucleotide sequence of Cauliflower Mosaïc Virus DNA. Cell 21: 285–294 (1980).

    Article  PubMed  Google Scholar 

  14. Gatignol A, Durand H, Tiraby G: Bleomycin resistance conferred by a drug binding protein. FEBS Lett 230: 171–175 (1988).

    Article  PubMed  Google Scholar 

  15. Genilloud O, Garrido MC, Moreno F: The transposon Tn5 carries a bleomycin resistance determinant. Gene 32: 225–233 (1984).

    Article  PubMed  Google Scholar 

  16. Grimwade JE, Cason EB, Beerman TA: Site specificity of bleomycin cleavage in purified and intracellular simian virus 40 DNA. Nucleic Acids Res 15: 6315–6329 (1987).

    PubMed  Google Scholar 

  17. Guerche P, Bellini C, LeMoullec JM, Caboche M: Use of a transient expression assay for the optimization of direct gene transfer into tobacco mesophyll protoplasts by electroporation. Biochimie 69: 621–628 (1987).

    Article  PubMed  Google Scholar 

  18. Hayford MB, Medford JI, Hoffman NL, Rogers SG, Klee HJ: Development of a plant transformation selection system based on expression of genes encoding gentamicin acetyltransferases. Plant Physiol 86: 1216–1222 (1988).

    Google Scholar 

  19. Herrera-Estrella L, DeBlock M, Messens E, Hernalsteens JP, VanMontagu M, Schell J: Chimaeric genes as dominant selectable markers in plant cells. EMBO J 2: 987–995 (1983).

    Google Scholar 

  20. Herrera-Estrella L, Depicker A, VanMontagu M, Schell J: Expression of chimaeric genes transferred into plant cells using a Ti plasmid derived vector. Nature 303: 209–213 (1983).

    Google Scholar 

  21. Hille J, Verheggen F, Roelvink P, Franssen H, VanKammen A, Zabel P: Bleomycin resistance: a new dominant selectable marker for plant cell transformation. Plant Mol Biol 7: 171–176 (1986).

    Google Scholar 

  22. Hoekema A, Hirsch PR, Hooykass PJJ, Schilperoort RA: A binary plant vector strategy based on separation of the Vir- and T-region of the Agrobacterium tumefaciens Ti-plasmid. Nature 303: 179–180 (1980).

    Google Scholar 

  23. Horsch RB, Fry JE, Hoffmann NL, Walbroth N, Eichholtz D, Rogers SG, Fraley RT: A simple and general method for transferring genes into plants. Science 227: 1229–1331 (1985).

    Google Scholar 

  24. Jones JDG, Svab Z, Harper EC, Hurwitz CD, Maliga P: A dominant nuclear streptomycin resistance marker for plant cell transformation. Mol Gen Genet 210: 86–91 (1987).

    Google Scholar 

  25. Krens FA, Molendijk L, Wullems GJ, Schilperoort RA: In vitro transformation of plant protoplasts with Ti-plasmid DNA. Nature 296: 72–74 (1982).

    Google Scholar 

  26. Kross J, Heuner WD, Hecht SM, Haseltine WA: Specificity of deoxyribonucleic acid cleavage by bleomycin, phleomycin and tallysomycin. Biochemistry 21: 4310–4318 (1982).

    PubMed  Google Scholar 

  27. Lörz H, Baker B, Schell J: Gene transfer to cereal cells mediated by protoplast transformation. Mol Gen Genet 199: 178–182 (1985).

    Google Scholar 

  28. Maniatis T, Fritsch E, Sambrook J: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).

    Google Scholar 

  29. Messing J: New M13 vectors for cloning. In: Methods in Enzymology 101, pp. 20–78. Academic Press, New York (1983).

    Google Scholar 

  30. Mulsant P, Gatignol A, Dalens M, Tiraby G: Phleomycin resistance as a dominant selectable marker in CHO cells. Som Cell Mol genet 14: 243–252 (1988).

    PubMed  Google Scholar 

  31. Paszkowski J, Shillito RD, Saul M, Mandak V, Hohn T, Hohn B, Potrykus I: Direct gene transfer to plants. EMBO J 3: 2717–2722 (1984).

    Google Scholar 

  32. Potrykus I, Saul MW, Petruska J, Paszkowski J, Shillito RD: Direct gene transfer to cells of a graminaceous monocot. Mol Gen Genet 199: 178–182 (1985).

    Google Scholar 

  33. Sanders PR, Winter JA, Barnason AR, Rogers SG, Fraley RT: Comparison of cauliflower mosaïc virus 35S and nopaline synthase promoters in transgenic plants. Nucleic Acids Res 15: 1543–1558 (1987).

    PubMed  Google Scholar 

  34. Twigg AJ, Sheratt D: Trans-complementable copy-number mutants of plasmid Col EI. Nature 283: 216–218 (1980).

    PubMed  Google Scholar 

  35. Umezawa H: Advances in bleomycin studies. In: Hecht SM (ed) Bleomycin, Biochemical and Biological Aspects, pp. 24–36. Springer-Verlag, New York (1979).

    Google Scholar 

  36. Van denElzen PJM, Townsend J, Lee KY, Bedbrook JR: A chimaeric hygromycin resistance gene as a selectable marker in plant cells. Plant Mol Biol 5: 299–302 (1985).

    Google Scholar 

  37. Waldron C, Murphy EB, Roberts JL, Gustafson GD, Armour SL, Malcom SK: Resistance to hygromycin B. A new marker for plant transformation studies. Plant Mol Biol 5: 103–108 (1985).

    Google Scholar 

  38. Zambryski P, Joos H, Genetello C, Leemans J, VanMontagu M, Schell J: Ti plasmid vector for introduction of DNA into plant cells without alteration of their normal regeneration capacity. EMBO J 2: 2143–2150 (1983).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. BIOSEM, Laboratoire de biologie moléculaire et cellulaire, Groupe LIMAGRAIN, Campus Universitaire des Cézeaux, 24, avenue des Landais, 63170, Aubiere, France

    Pascual Perez, Jean Kallerhoff & Joël Perret

  2. Laboratoire de microbiologie et génétique appliquées du CNRS, C.R.B.G.C., Université Paul Sabatier, 118, route de Narbonne, 31062, Toulouse Cédex, France

    Gérard Tiraby

Authors
  1. Pascual Perez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Gérard Tiraby
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jean Kallerhoff
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Joël Perret
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Perez, P., Tiraby, G., Kallerhoff, J. et al. Phleomycin resistance as a dominant selectable marker for plant cell transformation. Plant Mol Biol 13, 365–373 (1989). https://doi.org/10.1007/BF00015548

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00015548

Key words

Search

Navigation