title: Quantitative assay of photoinduced DNA strand breaks by real-time PCR.
abstract: Real-time PCR (qPCR) - a modern methodology primarily used for studying gene expression has been employed for the quantitative assay of an important class of DNA damage - single strand breaks. These DNA lesions which may lead to highly cytotoxic double strand breaks were quantified in a model system where double stranded DNA was sensitized to UV photons by labeling with 5-bromo-2'-deoxyuridine. The amount of breaks formed due to irradiation with several doses of 320nm photons was assayed by two independent methods: LC-MS and qPCR. A very good agreement between the relative damage measured by the two completely different analytical tools proves the applicability of qPCR for the quantitative analysis of SSBs. Our results suggest that the popularity of the hitherto underestimated though accurate and site-specific technique of real-time PCR may increase in future DNA damage studies.
authors.affiliation: Faculty of Chemistry, University of Gdańsk, Wita Stwosza 63, 80-308 Gdańsk, Poland.
authors.forename: Justyna
authors.initials: J
authors.lastname: Wiczk
authors.affiliation: Faculty of Chemistry, University of Gdańsk, Wita Stwosza 63, 80-308 Gdańsk, Poland.
authors.forename: Kinga
authors.initials: K
authors.lastname: Westphal
authors.affiliation: Faculty of Chemistry, University of Gdańsk, Wita Stwosza 63, 80-308 Gdańsk, Poland. Electronic address: janusz.rak@ug.edu.pl.
authors.forename: Janusz
authors.initials: J
authors.lastname: Rak
mesh_terms: D001973:Bromodeoxyuridine / Q000494:pharmacology; D002853:Chromatography, Liquid; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects / Q000528:radiation effects; D013058:Mass Spectrometry; D017319:Photosensitizing Agents / Q000494:pharmacology; D060888:Real-Time Polymerase Chain Reaction; D014466:Ultraviolet Rays / Q000009:adverse effects
chemical_list: D017319:Photosensitizing Agents; D001973:Bromodeoxyuridine
keywords: Mass spectrometry; Photodamage; Photosensitizing nucleobases; Real-time PCR; Single strand breaks