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Promoter methylation and progressive transgene inactivation inArabidopsis

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Abstract

Agrobacterium-transformedArabidopsis plants were generated and the stability of their T-DNA-encoded resistance to kanamycin was examined. Of seven families, each homozygous for a single insertion event, two showed progressive inactivation of resistance over four generations of inbreeding. Loss of resistance was associated with methylation of anSst II site in thenos promoter of the kanamycin resistance gene. Treatment of plant roots from inactive lines with the demethylating agent 5-azacytidine restored the ability of such lines to form callus on kanamycin-containing media. These observations are consistent with the view that methylation is a factor in the progressive inactivation of transgenes inArabidopsis.

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References

  1. Amasino RM, Powell ALT, Gordon MP: Changes in T-DNA methylation and expression are associated with phenotypic variation and plant regeneration in a crown gall tumor line. Mol Gen Genet 197: 437–446 (1984).

    PubMed  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. An G, Ebert PR, Yi BY, Choi CH: Both TATA box and upstream regions are required for nopaline synthase promoter activity in transformed tobacco cells. Mol Gen Genet 203: 245–250 (1986).

    Article  Google Scholar 

  4. Beck E, Ludwig G, Auerswald EA, Reiss B, Schaller H: Nucleotide sequence and exact localization of the neomycin phosphotransferase gene from transposon Tn 5. Gene 19: 327–336 (1982).

    Article  PubMed  Google Scholar 

  5. Bezdek M, Koulalová B, Brzobohaty B, Vyskot B: 5-Azacytidine-induced hypomethylation of tobacco HRS60 tandem DNA repeats in tissue culture. Planta 184: 487–490 (1991).

    Google Scholar 

  6. Budar F, Thia-Toong L, van Montagu M, Hernalsteens JP:Agrobacterium-mediated gene transfer results mainly in transgenic plants transmitting T-DNA as a single Mendelian factor. Genetics 114: 303–313 (1986).

    Google Scholar 

  7. Deroles SC, Gardner RC: Expression and inheritance of kanamycin resistance in a large number of transgenic petunias generated byAgrobacterium-mediated transformation. Plant Mol Biol 11: 355–364 (1988).

    Google Scholar 

  8. Flavell RB, O'Dell M: Regulation of cytosine methylation of ribosomal DNA and nucleolus organizer expression in wheat. J. Mol Biol 204: 523–534 (1988).

    Article  PubMed  Google Scholar 

  9. Gruenbaum Y, Naveh-Many T, Cedar H, Razin A: Sequence specificity of methylation in higher plant DNA. Nature 292: 860–862 (1981).

    PubMed  Google Scholar 

  10. Heberle-Bors E, Charvat B, Thompson D, Schernthaner JP, Barta A, Matzke AJM, Matzke MA: Genetic analysis of T-DNA insertions into the tobacco genome. Plant Cell Rep 7: 571–574 (1988).

    Article  Google Scholar 

  11. Hepburn AG, Clarke LE, Pearson L, White J: The role of cytosine methylation in the control of nopaline synthase gene expression in a plant tumor. J Mol Appl Genet 2: 315–329 (1983).

    PubMed  Google Scholar 

  12. Heslop-Harrison JS: Gene expression and parental dominance in hybrid plants. Dev Supp: 21–28 (1990).

  13. Hobbs SLA, Kpodar P, DeLong CMO: The effect of T-DNA copy number, position and methylation on reporter gene expression in tobacco transformants. Plant Mol Biol 15: 851–864 (1990).

    PubMed  Google Scholar 

  14. Jones JDG, Gilbert DE, Grady KL, Jorgensen RA: T-DNA structure and gene expression in petunia plants transformed byAgrobacterium tumefaciens C58 derivatives. Mol Gen Genet 207: 478–485 (1987).

    Article  Google Scholar 

  15. Linn F, Heidmann I, Saedler H, Meyer P: Epigenetic changes in the expression of the maize A1 gene inPetunia hybrida: Role of numbers of integrated gene copies and state of methylation. Mol Gen Genet 222: 329–336 (1990).

    PubMed  Google Scholar 

  16. Matzke MA, Primig M, Trnovsky J, Matzke AJM: Reversible methylation and inactivation of marker genes in sequentially transformed tobacco plants. EMBO J 8: 643–649 (1989).

    Google Scholar 

  17. Matzke MA, Matzke AJM: Gene interactions and epigenetic variation in transgenic plants. Devel Genet 11: 214–233 (1990).

    Google Scholar 

  18. Matzke MA, Matzke AJM: Differential inactivation and methylation of a transgene in plants by two suppressor loci containing homologous sequences. Plant Mol Biol 16: 821–830 (1991).

    Article  PubMed  Google Scholar 

  19. Murashige T, Skoog F: A revised medium for rapid growth and biossays with tobacco tissue cultures. Physiol Plant 15: 473–497 (1962).

    Google Scholar 

  20. Murray MG, Thompson WF: Rapid isolation of high molecular weight plant DNA. Nucl Acid Res 8: 4321–4325 (1980).

    Google Scholar 

  21. Peach C, Velten J: Transgene expression variability (position effect) of CAT and GUS reporter genes driven by linked divergent T-DNA promoters. Plant Mol Biol 17: 49–60 (1991).

    PubMed  Google Scholar 

  22. Peerbolte R, Leenhouts K, Hooykaas-van Slogteren GMS, Wullems GJ, Schilperoort RA: Clones from a shooty tobacco crown-gall tumor II: irregular T-DNA structures and organization, T-DNA methylation and conditional expression of opine genes. Plant Mol Biol 7: 285–299 (1986).

    Google Scholar 

  23. Potrykus I, Paszkowski R, Shillito RD, Saul MW: Direct gene transfer to plants. In: Hohn Th, Schell J (eds) Plant Gene Research: Plant DNA Infectious Agents, pp. 229–247. Springer-Verlag, Vienna/New York (1987).

    Google Scholar 

  24. Pruitt RE, Meyerowitz EM: Characterization of the genome ofArabidopsis thaliana. J Mol Biol 187: 169–183 (1986).

    PubMed  Google Scholar 

  25. Scheid OM, Paszkowski J, Potrykus I: Reversible inactivation of a transgene inArabidopsis thaliana. Mol Gen Genet 228: 104–112 (1991).

    PubMed  Google Scholar 

  26. Schmidt R, Willmitzer L. High efficiencyAgrobacterium tumefaciens-mediated transformation ofArabidopsis thaliana leaf and cotyledon explants. Plant Cell Rep 7: 583–586 (1988).

    Article  Google Scholar 

  27. van Slogteren GMS, Hooykaas PJJ, Schilperoort RA: Silent T-DNA genes in plant lines transformed byAgrobacterium tumefaciens are activated by grafting and by 5-azacytidine treatment. Plant Mol Biol 3: 333–336 (1984).

    Google Scholar 

  28. Topping JF, Wei W, Lindsey K: Functional tagging of regulatory elements in the plant genome. Development 112: 1009–1019 (1991).

    PubMed  Google Scholar 

  29. Valvekens D, van Montagu M, van Lijsebettens M:Agrobacterium tumefaciens-mediated transformation ofArabidopsis thaliana root explants by using kanamycin selection. Proc Natl Acad Sci US 85: 5536–5540 (1988).

    Google Scholar 

  30. Vyskot B, Brzobohaty B, Karlovska L, Bezdek M: Structural and functional stability of foreign genes in transgenic tobacco plants. Folia Biol (Praha) 35: 360–372 (1989).

    Google Scholar 

  31. Weber H, Ziechmann C, Graessmann A:In vitro DNA methylation inhibits gene expression in transgenic tobacco. EMBO J 9: 4409–4415 (1990).

    PubMed  Google Scholar 

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Author notes

  1. Nigel J. Kilby

    Present address: Institute of Biotechnology, University of Cambridge, Tennis Court Road, CB2 1QT, Cambridge, UK

  2. H. M. Ottoline Leyser

    Present address: Department of Biology, Indiana University, Jordan Hall 142, 47401, Bloomington, IN, USA

Authors and Affiliations

  1. Department of Genetics, University of Cambridge, CB2 3EH, Cambridge, UK

    Nigel J. Kilby, H. M. Ottoline Leyser & Ian J. Furner

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  1. Nigel J. Kilby
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  2. H. M. Ottoline Leyser
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  3. Ian J. Furner
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Kilby, N.J., Leyser, H.M.O. & Furner, I.J. Promoter methylation and progressive transgene inactivation inArabidopsis . Plant Mol Biol 20, 103–112 (1992). https://doi.org/10.1007/BF00029153

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  • DOI: https://doi.org/10.1007/BF00029153

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