High-Resolution Characterization of DNA/Protein Complexes in Living Bacteria

Authors

Justin P. Peters, Mayo Clinic Graduate School of Biomedical SciencesFollow
Nicole A. Becker, Mayo Clinic Graduate School of Biomedical SciencesFollow
L. James Maher, Mayo Clinic Graduate School of Biomedical SciencesFollow

Document Type

Article

Keywords

Architectural proteins, Chromatin endogenous cleavage (ChEC), Chromatin immunoprecipitation (ChIP), High-resolution mapping, Lac repression loop, Ligation-mediated PCR (LM-PCR), Phage lambda exonuclease, Polymerase chain reaction (PCR), Southern blot

Journal/Book/Conference Title

Methods in Molecular Biology

Volume

1837

First Page

95

Last Page

115

Abstract

The occurrence of DNA looping is ubiquitous. This process plays a well-documented role in the regulation of prokaryotic gene expression, such as the Escherichia coli lactose (lac) operon. Here, we present two complementary methods for high-resolution in vivo detection of DNA/protein binding within the bacterial nucleoid by using either chromatin immunoprecipitation combined with phage λ exonuclease digestion (ChIP-exo) or chromatin endogenous cleavage (ChEC), coupled with ligation-mediated polymerase chain reaction (LM-PCR) and Southern blot analysis. As an example we apply these in vivo protein-mapping methods to E. coli to show direct binding of architectural proteins in the Lac repressor-mediated DNA repression loop.

Department

Department of Chemistry and Biochemistry

Original Publication Date

1-1-2018

DOI of published version

10.1007/978-1-4939-8675-0_6

Recommended Citation

Peters, Justin P.; Becker, Nicole A.; and Maher, L. James, "High-Resolution Characterization of DNA/Protein Complexes in Living Bacteria" (2018). Faculty Publications. 6236.
https://scholarworks.uni.edu/facpub/6236