Abstract
The ability to detect and measure DNA single-strand breaks has been the aim of numerous assays developed to assess genotoxicity. These methods often rely on alkaline conditions to denature DNA. However, alkaline treatment of DNA also introduces artifactual SSBs through the cleavage of alkali-labile sites resulting in confounded data. Here, we describe a modified alkaline gel electrophoresis assay coupled with a neutral O-hydroxylamine to obtain the measurement of true SSB formation.
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References
Georgiou C, Papapostolou I, Grizntzalis K (2009) Protocol for the quantitative assessment of DNA concentration and damage (fragmentation and nicks). Nat Protoc 4:125–131
Anderson D, Plewa MJ (1998) The international comet assay workshop. Mutagenesis 13:67–73
Drouin R, Gao S, Homquist GP (1996) Agarose gel electrophoresis for DNA damage analysis. In: Pfeifer GP (ed) Technologies for detection of DNA damage and mutations. Plenum Press, New York, pp 37–43
Nakamura J et al (1998) Highly sensitive apurinic/apyrimidinic site assay can detect spontaneous and chemically induced depurination under physiological conditions. Cancer Res 58:222–225
Nakamura J, Swenberg JA (1999) Endogenous apurinic/apyrimidinic sites in genomic DNA of mammalian tissues. Cancer Res 59:2522–2526
Horton JK et al (2000) Protection against methylation-induced cytotoxicity by DNA polymerase beta-dependent long patch base excision repair. J Biol Chem 275:2211–2218
Acknowledgements
This work was supported by National Institute of Environmental Health Science grants T32-ES07126, P42-ES05948, and P30-ES10126 and National Cancer Institute grant T32-CA72319.
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© 2012 Springer Science+Business Media New York
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Luke, A.M., Nakamura, J. (2012). O-Hydroxylamine-Coupled Alkaline Gel Electrophoresis Assay for the Detection and Measurement of DNA Single-Strand Breaks. In: Bjergbæk, L. (eds) DNA Repair Protocols. Methods in Molecular Biology, vol 920. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-998-3_21
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DOI: https://doi.org/10.1007/978-1-61779-998-3_21
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Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-61779-997-6
Online ISBN: 978-1-61779-998-3
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