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. 1981 Jul;78(7):4344–4348. doi: 10.1073/pnas.78.7.4344

Differential expression of crown gall tumor markers in transformants obtained after in vitro Agrobacterium tumefaciens-induced transformation of cell wall regenerating protoplasts derived from Nicotiana tabacum

George J Wullems 1, Lucy Molendijk 1, Gert Ooms 1, Robbert A Schilperoort 1
PMCID: PMC319786  PMID: 16593059

Abstract

To obtain transformation of plant cells, we incubated 3-day-old cell wall-regenerating protoplasts from tobacco with Agrobacterium tumefaciens harboring tumor-inducing plasmids. Putative transformed tobacco cells were selected by phytohormone autotrophic growth and were shown to be transformed by the detection of the tumor cell specific enzymes lysopine dehydrogenase or nopaline dehydrogenase. This was substantiated by the detection, in transformed tumor tissues, of DNA sequences homologous to sequences in the tumor-inducing plasmid. Segregation of tumor markers has been observed among the transformants and it is suggested that this happened during the initiation of the transformation. The stable character of the transformed state was shown by the retention of tumor markers in subcloning of primary transformants under nonselective conditions. Suppression of the neoplastic state of transformants could take place, resulting in the development of transformed shoots. Indications were obtained for the inheritance of tumor markers through meiosis from seedlings obtained from seeds of flowering transformed plants that still expressed nopaline synthesis.

Keywords: tumor-inducing plasmids, opine synthesis, shoot regeneration, DNA-DNA hybridization

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bomhoff G., Klapwijk P. M., Kester H. C., Schilperoort R. A., Hernalsteens J. P., Schell J. Octopine and nopaline synthesis and breakdown genetically controlled by a plasmid of Agrobacterium tumefaciens. Mol Gen Genet. 1976 May 7;145(2):177–181. doi: 10.1007/BF00269591. [DOI] [PubMed] [Google Scholar]
  2. Braun A. C. Plant tumors. Biochim Biophys Acta. 1978 Oct 27;516(2):167–191. doi: 10.1016/0304-419x(78)90007-0. [DOI] [PubMed] [Google Scholar]
  3. Chilton M. D., Drummond M. H., Merio D. J., Sciaky D., Montoya A. L., Gordon M. P., Nester E. W. Stable incorporation of plasmid DNA into higher plant cells: the molecular basis of crown gall tumorigenesis. Cell. 1977 Jun;11(2):263–271. doi: 10.1016/0092-8674(77)90043-5. [DOI] [PubMed] [Google Scholar]
  4. Chilton M. D., Drummond M. H., Merio D. J., Sciaky D., Montoya A. L., Gordon M. P., Nester E. W. Stable incorporation of plasmid DNA into higher plant cells: the molecular basis of crown gall tumorigenesis. Cell. 1977 Jun;11(2):263–271. doi: 10.1016/0092-8674(77)90043-5. [DOI] [PubMed] [Google Scholar]
  5. Chilton M. D., Saiki R. K., Yadav N., Gordon M. P., Quetier F. T-DNA from Agrobacterium Ti plasmid is in the nuclear DNA fraction of crown gall tumor cells. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4060–4064. doi: 10.1073/pnas.77.7.4060. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hooykaas P. J., den Dulk-Ras H., Ooms G., Schilperoort R. A. Interactions between octopine and nopaline plasmids in Agrobacterium tumefaciens. J Bacteriol. 1980 Sep;143(3):1295–1306. doi: 10.1128/jb.143.3.1295-1306.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Klapwijk P. M., Scheulderman T., Schilperoort R. A. Coordinated regulation of octopine degradation and conjugative transfer of Ti plasmids in Agrobacterium tumefaciens: evidence for a common regulatory gene and separate operons. J Bacteriol. 1978 Nov;136(2):775–785. doi: 10.1128/jb.136.2.775-785.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Klapwijk P. M., van Breukelen J., Korevaar K., Ooms G., Schilperoort R. A. Transposition of Tn904 encoding streptomycin resistance into the octopine Ti plasmid of Agrobacterium tumefaciens. J Bacteriol. 1980 Jan;141(1):129–136. doi: 10.1128/jb.141.1.129-136.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Koekman B. P., Ooms G., Klapwijk P. M., Schilperoort R. A. Genetic map of an octopine TI-plasmid. Plasmid. 1979 Jul;2(3):347–357. doi: 10.1016/0147-619x(79)90018-0. [DOI] [PubMed] [Google Scholar]
  10. Maliga P., Sz-Breznovits A., Márton L. Streptomycin-resistant plants from callus culture of haploid tobacco. Nat New Biol. 1973 Jul 4;244(131):29–30. doi: 10.1038/newbio244029a0. [DOI] [PubMed] [Google Scholar]
  11. McPherson J. C., Nester E. W., Gordon M. P. Proteins encoded by Agrobacterium tumefaciens Ti plasmid DNA (T-DNA) in crown gall tumors. Proc Natl Acad Sci U S A. 1980 May;77(5):2666–2670. doi: 10.1073/pnas.77.5.2666. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ooms G., Klapwijk P. M., Poulis J. A., Schilperoort R. A. Characterization of Tn904 insertions in octopine Ti plasmid mutants of Agrobacterium tumefaciens. J Bacteriol. 1980 Oct;144(1):82–91. doi: 10.1128/jb.144.1.82-91.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Otten L. A., Schilperoort R. A. A rapid micro scale method for the detection of lysopine and nopaline dehydrogenase activities. Biochim Biophys Acta. 1978 Dec 8;527(2):497–500. doi: 10.1016/0005-2744(78)90363-7. [DOI] [PubMed] [Google Scholar]
  14. Thomashow M. F., Nutter R., Montoya A. L., Gordon M. P., Nester E. W. Integration and organization of Ti plasmid sequences in crown gall tumors. Cell. 1980 Mar;19(3):729–739. doi: 10.1016/s0092-8674(80)80049-3. [DOI] [PubMed] [Google Scholar]

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