Quantitation of chemically induced DNA strand breaks in human cells via an alkaline unwinding assay
Analytical BiochemistryPublished 1 February 1985
Florian Daniel, Dorothee Haas, SCOTT PYLE
Citations87
SJR quartileQ3
SJR score0.50
SNIP0.63
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TL;DR
A correlation was shown between mammalian DNA strand-breaking potential (as measured in this procedure) and the propensity of these chemicals to revert Salmonella typhimurium TA100.
Abstract
DNA strand breaks induced in human CCRF-CEM cells by electrophilic chemicals (carcinogens/mutagens) can be readily quantitated via a facile alkaline unwinding assay. This procedure estimates the number of chemically induced DNA strand breaks on the basis of the percentage DNA converted from double-stranded to single-stranded form during an exposure to the alkaline unwinding conditions. The assay is based on the assumption that each strand break serves as a strand unwinding point during the alkaline denaturation. The extent of strand separation can be standardized with respect to the initial level of induced strand breaks by the use of X-rays, which produce known levels of DNA strand breaks per rad in mammalian cells. Subsequent to the alkaline exposure, the single- and double-stranded DNA were separated by use of thermostated hydroxylapatite columns (60 degrees C), and the DNA was quantitated via a fluorescence assay (Hoechst 33258 compound). A correlation was shown between mammalian DNA strand-breaking potential (as measured in this procedure) and the propensity of these chemicals to revert Salmonella typhimurium TA100.
Keywords
Biochemistry, Genetics and Molecular Biology
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