Selective detection and quantification of oxidized abasic lesions in DNA

The C4′-oxidized abasic site (C4-AP) and 1,4-dioxobutane (DOB) are oxidized abasic lesions that are produced by a variety of DNA damaging agents. For instance, C4-AP accounts for∼40% of the lesions produced by the antitumor agent bleomycin. These lesions exhibit interesting and unique biochemical and biological effects in DNA. We have developed a simple, selective method for detecting femtomole quantities of C4-AP and DOB in DNA. The method takes advantage of the selective reactivity of the 1,4-dicarbonyl groups in the lesions with a biotinylated probe. Pretreatment of the DNA sample with DNA polymerase β enables one to distinguish between C4-AP and DOB. The adducts are quantified using a fluorescent assay. This is the first method for quantifying these lesions that does not require degradation of the biopolymer. It will be useful for analyzing the levels of these oxidized abasic lesions in DNA.