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Comet sensitivity in assessing DNA damage and repair in different cell cycle stages

Biomedical Sciences Research Institute Computer Science Research Institute Environmental Sciences Research Institute Nanotechnology & Advanced Materials Research Institute

McArt, DG, McKerr, George, Saetzler, Kurt, Howard, Vyvyan, Downes, Stephen and Wasson, GR (2010) Comet sensitivity in assessing DNA damage and repair in different cell cycle stages. Mutagenesis, 25 (3). p. 299. [Journal article]

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URL: http://dx.doi.org/10.1093/mutage/geq006

DOI: doi:10.1093/mutage/geq006

Abstract

The comet assay is a sensitive tool for estimation of DNA damage and repair at the cellular level, requiring only a very small number of cells. In comparing the levels of damage or repair in different cell samples, it is possible that small experimental effects could be confounded by different cell cycle states in the samples examined, if sensitivity to DNA damage, and repair capacity, varies with the cell cycle. We assessed this by arresting HeLa cells in various cell cycle stages and then exposing them to ionizing radiation. Unirradiated cells demonstrated significant differences in strand break levels measured by the comet assay (predominantly single-strand breaks) at different cell cycle stages, increasing from G1 into S and falling again in G2. Over and above this variation in endogenous strand break levels, a significant difference in susceptibility to breaks induced by 3.5 Gy ionizing radiation was also evident in different cell cycle phases. Levels of induced DNA damage fluctuate throughout the cycle, with cells in G1 showing slightly lower levels of damage than an asynchronous population. Damage increases as cells progress through S phase before falling again towards the end of S phase and reaching lowest levels in M phase. The results from repair experiments (where cells were allowed to repair for 10 min after exposure to ionizing radiation) also showed differences throughout the cell cycle with G1-phase cells apparently being the most efficient at repair and M-phase cells the least efficient. We suggest, therefore, that in experiments where small differences in DNA damage and repair are to be investigated with the comet assay, it may be desirable to arrest cells in a specific stage of the cell cycle or to allow for differential cycle distribution.

Item Type:Journal article
Faculties and Schools:Faculty of Life and Health Sciences
Faculty of Life and Health Sciences > School of Biomedical Sciences
Research Institutes and Groups:Biomedical Sciences Research Institute
Biomedical Sciences Research Institute > Molecular Medicine
Biomedical Sciences Research Institute > Molecular Medicine > Neuroscience & Neurodegenerative Diseases
ID Code:16290
Deposited By:Dr Kurt Saetzler
Deposited On:03 Nov 2010 16:31
Last Modified:23 Jun 2011 16:15

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