Abstract
Single-molecule fluorescence microscopy has been used for decades to quantify macromolecular dynamics occurring in specimens that are in direct contact with a coverslip. This has permitted in vitro analysis of single-molecule motion in various biochemically reconstituted systems as well as in vivo studies of single-molecule motion on cell membranes. More recently, thanks to improvements in fluorescent tags and microscopes, it has been possible to follow individual molecules inside thicker specimens such as the nucleus of living cells. This has enabled a detailed description of the live-cell binding of nuclear proteins to DNA. In this protocol we describe a method to quantify intranuclear binding using single-molecule tracking (SMT).
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Acknowledgements
We are grateful to Drs. Tatiana Karpova and Tatsuya Morisaki for constructive feedback on the manuscript. DM is funded by a Marie Curie International Incoming Fellowship [Grant agreement: 27432].
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Mazza, D., Ganguly, S., McNally, J.G. (2013). Monitoring Dynamic Binding of Chromatin Proteins In Vivo by Single-Molecule Tracking. In: Shav-Tal, Y. (eds) Imaging Gene Expression. Methods in Molecular Biology, vol 1042. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-526-2_9
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DOI: https://doi.org/10.1007/978-1-62703-526-2_9
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