Abstract
Fluorescence microscopy has enabled the analysis of both the spatial distribution of DNA damage and its dynamics during the DNA damage response (DDR). Three microscopic techniques can be used to study the spatiotemporal dynamics of DNA damage. In the first part we describe how we determine the position of DNA double-strand breaks (DSBs) relative to the nuclear envelope. The second part describes how to quantify the co-localization of DNA DSBs with nuclear pore clusters, or other nuclear subcompartments. The final protocols describe methods for the quantification of locus mobility over time.
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Acknowledgement
We thank J. E. Haber for yeast strains and the Friedrich Miescher Institute Microscopy Facility for technical help. C.H. thanks the Marie Curie International program and JSPS Research Abroad program for fellowships. The Gasser laboratory thanks the Novartis Research Foundation, the Swiss National Science Foundation “Sinergia grant,” NCCR “Frontiers in Genetics,” and the Human Frontier Science Program (RGP0017).
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Horigome, C., Dion, V., Seeber, A., Gehlen, L.R., Gasser, S.M. (2015). Visualizing the Spatiotemporal Dynamics of DNA Damage in Budding Yeast. In: Oslowski, C. (eds) Stress Responses. Methods in Molecular Biology, vol 1292. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2522-3_6
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DOI: https://doi.org/10.1007/978-1-4939-2522-3_6
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