Visualization of chromatin dynamics by live cell microscopy using crispr/cas9 gene editing and anchor labeling

Abstract

The organization of the eukaryotic nucleus facilitates functional chromatin contacts which regulate gene transcription. Despite this being extensively studied through population-based chromatin contact mapping and microscopic observations in single cells, the spatiotemporal dynamics of chromatin behavior have largely remained elusive. The current methods to label and observe specific endogenous genomic loci in living cells have been challenging to implement and too invasive to biological processes. In this protocol, we describe the use of a recently developed DNA labelling strategy (ANCHOR) with CRISPR/Cas9 gene editing, to discreetly label genes for live cell imaging to study chromatin dynamics. Our approach improves on some of the fundamental shortfalls associated with current labelling strategies and has the potential for multiplexed observations.