Strand breaks in whole plasmid dna produced by the decay of (125)I in a triplex-forming oligonucleotide.
DNA strand breaks produced by the decay of (125)I positioned against a specific site in plasmid DNA via a triplex-forming oligonucleotide were studied both in the immediate vicinity of the site of the decay with a single nucleotide resolution and in the whole plasmid by measuring the percentages of supercoiled, open-circular and linear forms. The localized breaks are distributed within 10 bp in each direction from the decay site with maxima in both strands just opposite the (125)I-dC residue in the triplex-forming oligonucleotide. The distributions of breaks in the two DNA strands are almost symmetrical, in agreement with the geometry of the pyrimidine motif triplex. We found that about 25% of the double-strand breaks were located outside the 90-bp fragment containing the triplex-forming oligonucleotide binding sequence. The ratio of single- to double-strand breaks in the whole plasmid was 11 for bound triplex-forming oligonucleotide compared to 26 when the triplex-forming oligonucleotide was free in solution. The number of double-strand breaks per decay of (125)I was 0.46 for bound triplex-forming oligonucleotide and 0.17 for free triplex-forming oligonucleotide. Comparing the data on the localized damage and those for the whole plasmid, we concluded that, in addition to DNA breaks that are confined to a helical turn around the (125)I atom, the decay can produce breaks hundreds of base pairs away in the plasmid molecule. This linear plasmid molecule containing radiation-induced damage at a specific DNA site should be useful in studies of the molecular mechanisms of DNA repair.
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Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda Maryland 20854, USA. Irina@nmdpet.cc.nih.gov.
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D001483:Base Sequence; D004249:DNA Damage; D004260:DNA Repair; D019450:Genes, MDR; D006801:Humans; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes / Q000009:adverse effects*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry / Q000528:radiation effects*; D010957:Plasmids / Q000737:chemistry / Q000235:genetics / Q000528:radiation effects*
D007457:Iodine Radioisotopes; D009838:Oligodeoxyribonucleotides