CRISPR-dCas9 and sgRNA scaffolds enable dual-colour live imaging of satellite sequences and repeat-enriched individual loci
Year: 2016
Type: article
Abstract: Imaging systems that allow visualization of specific loci and nuclear structures are highly relevant for investigating how organizational changes within the nucleus play a role in regulating gene expression and other cellular processes. Here we present a live imaging system for targeted detection of genomic regions. Our approach involves generating chimaeric transcripts of viral RNAs (MS2 and PP7) and single-guide RNAs (sgRNAs), which when co-expressed with a cleavage-deficient Cas9 can recruit fluorescently tagged viral RNA-binding proteins (MCP and PCP) to specific genomic sites. This allows for rapid, stable, low-background visualization of target loci. We demonstrate the efficiency and flexibility of our method by simultaneously labelling major and minor satellite regions as well as two individual loci on mouse chromosome 12. This system provides a tool for dual-colour labelling, which is important for tracking the dynamics of chromatin interactions and for validating epigenetic processes identified in fixed cells.
Source: Nature Communications
DOI: 10.1038/ncomms11707
Institutions: New York University, NYU Langone Health
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $6,990
Awards: Enhancer Driven Gene Regulation During Lymphocyte Development, Molecular mechanisms of direct neuronal programming, A novel role for 53BP1 independent of damage that controls chromatin structure, Role Of Nuclear Organization In Protecting Genome Stability During Recombination, Cancer Center Support Grant
OpenAlex ID: W2401218948