A repair competition assay to assess recognition by human nucleotide excision repair.
We developed a competition assay to compare, in a quantitative manner, the ability of human nucleotide excision repair (NER) to recognise structurally different forms of DNA damage. This assay uses a NER substrate consisting of M13 double-stranded DNA with a single and uniquely located acetylaminofluorene (AAF) adduct, and measures the efficiency by which multiply damaged plasmid DNA competes for excision repair of the site-directed modification. To validate this assay, we tested competitor DNA containing defined numbers of either AAF adducts or UV radiation products. In both cases, repair of the site-directed NER substrate was inhibited in a damage-specific and dose-dependent manner. We then exploited this competition assay to determine the susceptibility of bulky adozelesin-DNA adducts to human NER.
Hess
M T
MT
Institute of Pharmacology and Toxicology, University of Zürich-Tierspital, Zürich, Switzerland.
Gunz
D
D
Naegeli
H
H
D001483:Base Sequence; D001572:Benzofurans; D001667:Binding, Competitive; D003509:Cyclohexanecarboxylic Acids; D053138:Cyclohexenes; D004249:DNA Damage; D004260:DNA Repair; D000080890:Duocarmycins; D006801:Humans; D007211:Indoles; D015336:Molecular Probe Techniques; D008969:Molecular Sequence Data
D001572:Benzofurans; D003509:Cyclohexanecarboxylic Acids; D053138:Cyclohexenes; D000080890:Duocarmycins; D007211:Indoles; C060090:adozelesin