Visualizing DNA damage and repair using single molecule super resolution microscopy

Sophie T B Morgan; Donna R Whelan; Ashley M Rozario

doi:10.1016/bs.mcb.2023.02.004

Visualizing DNA damage and repair using single molecule super resolution microscopy

Methods Cell Biol. 2024:182:237-245. doi: 10.1016/bs.mcb.2023.02.004. Epub 2023 Aug 21.

Authors

Sophie T B Morgan¹, Donna R Whelan¹, Ashley M Rozario²

Affiliations

¹ La Trobe Institute for Molecular Science, La Trobe Rural Health School, La Trobe University, Bendigo, VIC, Australia.
² La Trobe Institute for Molecular Science, La Trobe Rural Health School, La Trobe University, Bendigo, VIC, Australia. Electronic address: a.rozario@latrobe.edu.au.

PMID: 38359980
DOI: 10.1016/bs.mcb.2023.02.004

Abstract

Single molecule super resolution microscopy overcomes the diffraction limit by separating individual fluorophore emissions over time, resulting in spatial resolutions that are far superior to epifluorescence microscopy. This allows for DNA damage response (DDR) events to be investigated in greater detail. A variety of DNA damaging drugs can be used on S-phase synchronized immortalized cell lines alongside 5-ethynyl-2'-deoxyuridine (EdU) pulse labelling to ultimately visualize DNA repair pathways at distinct time points and quantify colocalizations between nascent DNA and immunolabeled DDR proteins. This chapter will outline super resolution microscopy assays to interrogate the spatiotemporal organization of DNA repair proteins at damaged foci during DDR events within immortalized cell lines.

Keywords: DNA damage response; DNA double strand break; Super resolution microscopy; dSTORM.

MeSH terms

Cell Line
DNA / genetics
DNA / metabolism
DNA Damage*
DNA Repair
Microscopy*
Single Molecule Imaging / methods

Substances

DNA