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High‐throughput single‐molecule studies of protein–DNA interactions
Author(s) -
Robison Aaron D.,
Finkelstein Ilya J.
Publication year - 2014
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2014.05.021
Subject(s) - fluorophore , nucleic acid , dna , fluorescence , computational biology , molecule , throughput , single molecule experiment , nanotechnology , small molecule , biophysics , chemistry , biology , computer science , biochemistry , physics , materials science , telecommunications , organic chemistry , quantum mechanics , wireless
Fluorescence and force‐based single‐molecule studies of protein–nucleic acid interactions continue to shed critical insights into many aspects of DNA and RNA processing. As single‐molecule assays are inherently low‐throughput, obtaining statistically relevant datasets remains a major challenge. Additionally, most fluorescence‐based single‐molecule particle‐tracking assays are limited to observing fluorescent proteins that are in the low‐nanomolar range, as spurious background signals predominate at higher fluorophore concentrations. These technical limitations have traditionally limited the types of questions that could be addressed via single‐molecule methods. In this review, we describe new approaches for high‐throughput and high‐concentration single‐molecule biochemical studies. We conclude with a discussion of outstanding challenges for the single‐molecule biologist and how these challenges can be tackled to further approach the biochemical complexity of the cell.