Petunia plants escape from negative selection against a transgene by silencing the foreign DNA via methylation View Full Text


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Article Info

DATE

1992-05

AUTHORS

Suzy Renckens, Henri De Greve, Marc Van Montagu, Jean-Pierre Hernalsteens

ABSTRACT

TransgenicPetunia hybrida clones harbouring the T-DNA gene2 ofAgrobacterium tumefaciens were used to test a strategy for the trapping of plant transposable elements. In thePetunia line used, floral variegation is due to the presence of the non-autonomous transposable elementdTph1 at theAn1 locus. The gene2 product converts the auxin precursor indole-3-acetamide and its analogue 1-naphthalene acetamide into the active auxins indole-3-acetic acid and 1-naphthalene acetic acid. Plant cells that express gene2 can use a low concentration of the precursors as auxins and become sensitive to the toxicity of high concentrations of these compounds. By selecting protoplast-derived microcalli or seedlings able to grow on medium with high precursor concentrations, variant plants were obtained in which gene2 was no longer expressed. Southern analysis, using gene2-specific probes, revealed that in one variant the T-DNA was deleted. For 30 other variants no alteration in gene2 structure was observed, indicating that transposable element insertion was not responsible for the inactivation of gene2. Analysis with restriction enzymes allowing discrimination between methylated or non-methylated DNA sequences showed that the inactivated gene2 sequences were methylated. Addition of the in vivo methylation inhibitor 5-azacytidine to the medium led to reactivation of gene2 expression in some of the variants. These observations demonstrated that reversible DNA methylation was the main cause of silencing of gene2 in this system. More... »

PAGES

53-64

References to SciGraph publications

  • 1984-02. Genetic analysis of instability in Petunia hybrida in THEORETICAL AND APPLIED GENETICS
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  • 1983-09. The frequency and distribution of methylatable DNA sequences in leguminous plant protein coding genes in JOURNAL OF MOLECULAR EVOLUTION
  • 1992-02. Endogenous and environmental factors influence 35S promoter methylation of a maize A1 gene construct in transgenic petunia and its colour phenotype in MOLECULAR GENETICS AND GENOMICS
  • 1989-03. DNA methylation of embryogenic carrot cell cultures and its variations as caused by mutation, differentiation, hormones and hypomethylating drugs in THEORETICAL AND APPLIED GENETICS
  • 1984-11. Silent T-DNA genes in plant lines transformed by Agrobacterium tumefaciens are activated by grafting and by 5-azacytidine treatment in PLANT MOLECULAR BIOLOGY
  • 1990-11. DNA variation in tissue-culture-derived rice plants in THEORETICAL AND APPLIED GENETICS
  • 1984-04. Genetic analysis of the individual T-DNA genes of Agrobacterium tumefaciens; further evidence that two genes are involved in indole-3-acetic acid synthesis in MOLECULAR GENETICS AND GENOMICS
  • 1988-12. Genetic analysis of T-DNA insertions into the tobacco genome in PLANT CELL REPORTS
  • 1977-01. Lambdoid phages that simplify the recovery of in vitro recombinants in MOLECULAR GENETICS AND GENOMICS
  • 1983-09. A plant DNA minipreparation: Version II in PLANT MOLECULAR BIOLOGY REPORTER
  • 1989-04. Rapid induction of genomic demethylation and T-DNA gene expression in plant cells by 5-azacytosine derivatives in PLANT MOLECULAR BIOLOGY
  • 1990. Do We Understand Somaclonal Variation? in PROGRESS IN PLANT CELLULAR AND MOLECULAR BIOLOGY
  • 1990-07. Epigenetic changes in the expression of the maize A1 gene inPetunia hybrida: Role of numbers of integrated gene copies and state of methylation in MOLECULAR GENETICS AND GENOMICS
  • 1990-09. Predominance and tissue specificity of adenine methylation in rice in THEORETICAL AND APPLIED GENETICS
  • 1981-08. Sequence specificity of methylation in higher plant DNA in NATURE
  • 1980-06. Nutritional requirements of protoplast-derived, haploid tobacco cells grown at low cell densities in liquid medium in PLANTA
  • 1988-01. A negative selection scheme for tobacco protoplast-derived cells expressing the T-DNA gene 2 in PLANT CELL REPORTS
  • 1989-03. Molecular characterization of Mutator systems in maize embryogenic callus cultures indicates Mu element activity in vitro in THEORETICAL AND APPLIED GENETICS
  • 1991-08. Reversible inactivation of a transgene in Arabidopsis thaliana in MOLECULAR GENETICS AND GENOMICS
  • 1986-03. Evidence for transposition in Petunia in THEORETICAL AND APPLIED GENETICS
  • 1980. Biosynthesis and Metabolism of Plant Hormones in HORMONAL REGULATION OF DEVELOPMENT I
  • 1985-11. Genetic evidence that the tryptophan 2-mono-oxygenase gene of Pseudomonas savastonoi is functionally equivalent to one of the T-DNA genes involved in plant tumour formation by Agrobacterium tumefaciens in MOLECULAR GENETICS AND GENOMICS
  • 1988-05. Expression and inheritance of kanamycin resistance in a large number of transgenic petunias generated by Agrobacterium-mediated transformation in PLANT MOLECULAR BIOLOGY
  • 1988-03. DNA methylation and tissue-specific transcription of the storage protein genes of maize in PLANT MOLECULAR BIOLOGY
  • Identifiers

    URI

    http://scigraph.springernature.com/pub.10.1007/bf00587561

    DOI

    http://dx.doi.org/10.1007/bf00587561

    DIMENSIONS

    https://app.dimensions.ai/details/publication/pub.1009889609

    PUBMED

    https://www.ncbi.nlm.nih.gov/pubmed/1376407


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    48 schema:description TransgenicPetunia hybrida clones harbouring the T-DNA gene2 ofAgrobacterium tumefaciens were used to test a strategy for the trapping of plant transposable elements. In thePetunia line used, floral variegation is due to the presence of the non-autonomous transposable elementdTph1 at theAn1 locus. The gene2 product converts the auxin precursor indole-3-acetamide and its analogue 1-naphthalene acetamide into the active auxins indole-3-acetic acid and 1-naphthalene acetic acid. Plant cells that express gene2 can use a low concentration of the precursors as auxins and become sensitive to the toxicity of high concentrations of these compounds. By selecting protoplast-derived microcalli or seedlings able to grow on medium with high precursor concentrations, variant plants were obtained in which gene2 was no longer expressed. Southern analysis, using gene2-specific probes, revealed that in one variant the T-DNA was deleted. For 30 other variants no alteration in gene2 structure was observed, indicating that transposable element insertion was not responsible for the inactivation of gene2. Analysis with restriction enzymes allowing discrimination between methylated or non-methylated DNA sequences showed that the inactivated gene2 sequences were methylated. Addition of the in vivo methylation inhibitor 5-azacytidine to the medium led to reactivation of gene2 expression in some of the variants. These observations demonstrated that reversible DNA methylation was the main cause of silencing of gene2 in this system.
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