How to get Live Imaging of the Cellular Genome via CRISPR System?

The spatiotemporal organization of chromatin structure plays a critical role in regulating lineage-specific gene expression during cellular differentiation and embryonic development, via contributing to the functional output of the genome, which can be dynamically amplified or suppressed. However, there is little known about how the genomes being modified and how their structural organization regulating function output.

The CRISPR-Cas9 genome editing system consists of two parts: 1- a Caspase enzyme called 'Cas9' with catalytic activity that cuts DNA strands by hydrolyzing phosphodiester bonds, and 2-a single guide RNA (sgRNA) that directs Cas9 to target sites with complementarity to nucleotides of the sgRNA in order to edit these sequences. Using a catalytically dead Cas9 (dCas9) it is possible to repurpose the CRISPR-Cas9 system, for example, by fusing dCas9 to epigenetic enzymes for epigenome editing or by fusing dCas9 to the Green Fluorescent Protein (GFP) for imaging of the cellular genome. Thus, it is possible to use dCas9 to create a customizable DNA labeler that can be complemented with a fluorescence microscope and thus track multiple genomic loci in living cells.