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Tethered particle experiments use light microscopy to measure the position of a micrometer-sized bead tethered to a microscope slide via an approximately micrometer-length polymer, to infer the behavior of the invisible polymer. Currently, this method is used to measure rate constants of DNA loop formation and breakdown mediated by repressor protein that binds to the DNA. We report a new technique for measuring these rates using a modified hidden Markov analysis that directly incorporates the diffusive motion of the bead, which is an inherent complication of tethered particle motion because it occurs on a timescale between the sampling frequency and the looping time. We compare looping lifetimes found with our method, which are consistent over a range of sampling frequencies, to those obtained via the traditional threshold-crossing analysis, which vary depending on how the raw data are filtered in the time domain. Our method does not involve such filtering, and so can detect short-lived looping events and sudden changes in looping behavior.
Beausang, J. F., Zurla, C., Manzo, C., Dunlap, D., Finzi, L., & Nelson, P. C. (2007). DNA Looping Kinetics Analyzed Using Diffusive Hidden Markov Model. Biophysics Journal, 92 (8), L64-L66. http://dx.doi.org/10.1529/biophysj.107.104828
Date Posted: 01 May 2017
This document has been peer reviewed.