Abstract
Faithful mitochondrial DNA (mtDNA) replication is critical for the proper function of the oxidative phosphorylation system. Problems with mtDNA maintenance, such as replication stalling upon encountering DNA damage, impair this vital function and can potentially lead to disease. An in vitro reconstituted mtDNA replication system can be used to investigate how the mtDNA replisome deals with, for example, oxidatively or UV-damaged DNA. In this chapter, we provide a detailed protocol on how to study the bypass of different types of DNA damage using a rolling circle replication assay. The assay takes advantage of purified recombinant proteins and can be adapted to the examination of various aspects of mtDNA maintenance.
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Acknowledgments
We thank Andreas Berner for his technical input. We thank Prof. Peter Burgers for the experiments carried out in his laboratory (Washington University in St. Louis). This work was supported by the Knut and Alice Wallenberg Foundation, and the Swedish Research Council.
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Forslund, J.M.E., Stojkovič, G., Wanrooij, S. (2023). Rolling Circle Replication and Bypass of Damaged Nucleotides. In: Nicholls, T.J., Uhler, J.P., Falkenberg, M. (eds) Mitochondrial DNA. Methods in Molecular Biology, vol 2615. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2922-2_15
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DOI: https://doi.org/10.1007/978-1-0716-2922-2_15
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