Single-Molecule Real-Time Visualization of DNA Unwinding by CMG Helicase

In our lab, we are studying the molecular mechanisms of eukaryotic DNA replication. We want to understand the dynamics of genome duplication at the single-molecule level. We visualize DNA replication using multiple systems, including purified proteins and extract derived from Xenopus eggs.

Much of what we know about DNA replication has come from bulk experiments, cell imaging, and structural biology. These methods are really powerful, but they're often limited in what they can tell us about the dynamics or transient behaviors of individual proteins. This is what single-molecule techniques are really well-suited to studying, and it's why their use is growing in the field.

Our protocol for live visualization of DNA unwinding outlines a platform that can be built upon and modified to investigate different aspects of DNA replication. It can provide a better understanding of the molecular mechanism of CMG activity in the presence of chosen ribosome components and replicative stress. To understand the complex systems at play during DNA replication, we isolate key proteins and visualize their real-time behavior in vitro.

The insights we gain contribute to a more complete picture of DNA replication.