The complex binding dynamics between DNA and proteins are often obscured by ensemble averaging effects in conventional biochemical experiments. Single-molecule fluorescence methods are powerful tools to investigate DNA–protein interaction dynamics in real time. In this chapter, we focus on using single-molecule Förster Resonance Energy Transfer (smFRET) to probe the binding dynamics of individual proteins on single DNA molecules. We provide a detailed discussion of total internal reflection fluorescence (TIRF) instrument design, nucleic acid labeling with fluorophores, flow cell surface passivation, and data analysis methods.
CITATION STYLE
Chaurasiya, K. R., & Dame, R. T. (2018). Single molecule FRET analysis of DNA binding proteins. In Methods in Molecular Biology (Vol. 1665, pp. 217–239). Humana Press Inc. https://doi.org/10.1007/978-1-4939-7271-5_12
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