Visualizing DNA replication at the single-molecule level

Methods in Enzymology
Nathan A Tanner, Antoine M van Oijen

Abstract

Recent advances in single-molecule methodology have made it possible to study the dynamic behavior of individual enzymes and their interactions with other proteins in multiprotein complexes. Here, we describe newly developed methods to study the coordination of DNA unwinding, priming, and synthesis at the DNA-replication fork. The length of individual DNA molecules is used to measure the activity of single replisomes engaged in coordinated DNA replication. First, a tethered-particle technique is used to visualize the formation and release of replication loops. Second, a fluorescence imaging method provides a direct readout of replication rates and processivities from individual replisomes. The ability to directly observe transient reaction intermediates and characterize heterogeneous behavior makes these single-molecule approaches important new additions to the tools available to study DNA replication.

Citations

Jun 21, 2011·Genes & Development·Ignacio Tinoco, Ruben L Gonzalez
Sep 29, 2011·International Journal of Molecular Sciences·Andreas E Vasdekis, Gregoire P J Laporte
Mar 5, 2014·Proceedings of the National Academy of Sciences of the United States of America·Hylkje J GeertsemaAntoine M van Oijen
May 16, 2014·Proceedings of the National Academy of Sciences of the United States of America·James E KathJoseph J Loparo
Jul 7, 2016·Nucleic Acids Research·Kelly M SchermerhornAndrew F Gardner
Mar 29, 2011·The EMBO Journal·Nathan A TannerAntoine M van Oijen
Nov 10, 2018·PloS One·Tom E P Kimkes, Matthias Heinemann
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Sep 1, 2015·PLoS Genetics·Andrew RobinsonAntoine M van Oijen
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Apr 29, 2014·Biotechnology and Bioengineering·Samaneh Mashaghi, Antoine M van Oijen
Nov 16, 2019·Scientific Reports·Arash AhmadiBjørn Dalhus
Dec 21, 2018·Nature Communications·Arash AhmadiAlexander D Rowe
Apr 5, 2019·Biochemistry·Andrew A DrabekStephen C Blacklow

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