Nucleic Acids Research, Vol 27, Issue 21 4282-4290, Copyright © 1999 by Oxford University Press
K Mezhevaya, TA Winters and RD Neumann
A parallel binding motif 16mer triplex-forming oligonucleotide (TFO)
complementary to a polypurine-polypyrimidine target region near the 3'- end
of the SupF gene of plasmid pSP189 was labeled with [5-(125)I]dCMP at
position 15. Following triplex formation and decay accumulation,
radiation-induced site-specific double-strand breaks (DSBs) were produced
in the pSP189 SupF gene. Bulk damaged DNA and the isolated site-specific
DSB-containing DNA were separately transfected into human WI38VA13 cells
and allowed to repair prior to recovery and analysis of mutants. Bulk
damaged DNA had a relatively low mutation frequency of 2.7 x 10(-3). In
contrast, the isolated linear DNA containing site- specific DSBs had an
unusually high mutation frequency of 7.9 x 10(-1). This was nearly 300-fold
greater than that observed for the bulk damaged DNA mixture, and >1.5 x
10(4)-fold greater than background. The mutation spectra displayed a high
proportion of deletion mutants targeted to the(125)I binding position
within the SupF gene for both bulk damaged DNA and isolated linear DNA.
Both spectra were characterized by complex mutations with mixtures of
changes. However, mutations recovered from the linear site-specific
DSB-containing DNA presented a much higher proportion of complex deletion
mutations.
ARTICLES
Gene targeted DNA double-strand break induction by (125)I-labeled triplex-forming oligonucleotides is highly mutagenic following repair in human cells
Department of Nuclear Medicine, Warren Grant Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892, USA.
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