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Updated: Dec 11, 2025

Inducing a Site Specific Replication Blockage in E. coli Using a Fluorescent Repressor Operator System
Published on: August 21, 2016
The Analysis of Recombination-Dependent Processing of Blocked Replication Forks by Bidimensional Gel Electrophoresis
Karol Kramarz1,2,3, Anissia Ait Saada4, Sarah A E Lambert5,6,7
1Institut Curie, CNRS, UMR3348, PSL Research University, Orsay, France.
Abstract:
The perturbation of the DNA replication process is a threat to genome stability and is an underlying cause of cancer development and numerous human diseases. It has become central to understanding how stressed replication forks are processed to avoid their conversion into fragile and pathological DNA structures. The engineering of replication fork barriers (RFBs) to conditionally induce the arrest of a single replisome at a defined locus has made a tremendous impact in our understanding of replication fork processing. Applying the bidimensional gel electrophoresis (2DGE) technique to those site-specific RFBs allows the visualization of replication intermediates formed in response to replication fork arrest to investigate the mechanisms ensuring replication fork integrity. Here, we describe the 2DGE technique applied to the site-specific RTS1-RFB in Schizosaccharomyces pombe and explain how this approach allows the detection of arrested forks undergoing nascent strands resection.
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