Translesion DNA synthesis (TLS) is an evolutionarily conserved branch of the cellular DNA damage tolerance pathway that is often exploited by cancer cells to overcome therapy resistance. Here, we present a protocol to analyze endogenous TLS in single mammalian cells in the absence or presence of DNA damage. We describe steps for detecting chromatin-bound TLS factors, such as monoubiquitinated PCNA(mUb) and TLS DNA polymerases (pols) by flow cytometry. We then detail a procedure to detect their nuclear localization using immunofluorescence. For complete details on the use and execution of this protocol, please refer to Egger et al. (Cell Reports Methods, in press)..
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| http://dx.doi.org/10.1016/j.xpro.2023.102361 | DOI Listing |
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