Please use this identifier to cite or link to this item: http://hdl.handle.net/2440/131262
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Type: Journal article
Title: The loopometer: a quantitative in vivo assay for DNA-looping proteins
Author: Hao, N.
Sullivan, A.E.
Shearwin, K.E.
Dodd, I.B.
Citation: Nucleic Acids Research, 2021; 49(7):e39-1-e39-16
Publisher: Oxford University Press (OUP)
Issue Date: 2021
ISSN: 0305-1048
1362-4962
Statement of
Responsibility: 
Nan Hao, Adrienne E. Sullivan, Keith E. Shearwin and Ian B. Dodd
Abstract: Proteins that can bring together separate DNA sites, either on the same or on different DNA molecules, are critical for a variety of DNA-based processes. However, there are no general and technically simple assays to detect proteins capable of DNA looping in vivo nor to quantitate their in vivo looping efficiency. Here, we develop a quantitative in vivo assay for DNA-looping proteins in Escherichia coli that requires only basic DNA cloning techniques and a LacZ assay. The assay is based on loop assistance, where two binding sites for the candidate looping protein are inserted internally to a pair of operators for the E. coli LacI repressor. DNA looping between the sites shortens the effective distance between the lac operators, increasing LacI looping and strengthening its repression of a lacZ reporter gene. Analysis based on a general model for loop assistance enables quantitation of the strength of looping conferred by the protein and its binding sites. We use this ‘loopometer’ assay to measure DNA looping for a variety of bacterial and phage proteins.
Keywords: Nucleic Acid Conformation
Rights: © The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
RMID: 1000033902
DOI: 10.1093/nar/gkaa1284
Grant ID: http://purl.org/au-research/grants/nhmrc/GNT1100651
http://purl.org/au-research/grants/arc/DP160101450
Appears in Collections:Biochemistry publications

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