Rare de novo methylation within the transposable element activator (Ac) in transgenic tobacco plants

Transgenic SR1 tobacco plants that contained the maize transposable element Ac or deletion derivatives thereof were isolated. The DNA methylation patterns of the foreign DNA sequences were analysed with methylation-sensitive restriction enzymes. By this method we tested 87 cytosine residues whose methylation is known to inhibit restriction by the corresponding enzyme. Most of the restriction sites were cleavable and hence unmethylated in transgenic tobacco plants. There were only three restriction sites at which a fraction of the Ac sequences was methylated. A similar result was obtained with two inactive, internally or terminally deleted Ac sequences. In one of the deletion derivatives a single restriction site was completely methylated. All other sites were unmethylated. The complete Ac elements described in this report were present as single copies in the transgenic plants. Their activity was demonstrated by the presence of the element-specific transcript. The deletion derivatives did not transpose in the transgenic tobacco plants and were thus still linked to the sequences of the plasmid that was used for transformation. These adjacent sequences represent part of a chimaeric NPTII gene inactivated by the insertion of the Ac deletion derivative. All 16 restriction sites examined in this sequence were unmethylated.