ZFIN ID: ZDB-PUB-190604-2
Chemical reprogramming enhances homology-directed genome editing in zebrafish embryos
Aksoy, Y.A., Nguyen, D.T., Chow, S., Chung, R.S., Guillemin, G.J., Cole, N.J., Hesselson, D.
Date: 2019
Source: Communications biology   2: 198 (Journal)
Registered Authors: Aksoy, Yagiz, Chow, Sharon, Chung, Roger, Cole, Nicholas, Hesselson, Daniel
Keywords: CRISPR-Cas9 genome editing, Chemical biology
MeSH Terms:
  • Animals
  • CRISPR-Cas Systems*
  • Clustered Regularly Interspaced Short Palindromic Repeats
  • DNA Breaks, Double-Stranded
  • DNA End-Joining Repair
  • Gene Editing*
  • Genotype
  • Green Fluorescent Proteins/metabolism
  • RNA/metabolism
  • Recombinational DNA Repair
  • Zebrafish/embryology*
  • Zebrafish/genetics*
PubMed: 31149642 Full text @ Commun Biol
Precise genome editing is limited by the inefficiency of homology-directed repair (HDR) compared to the non-homologous end-joining (NHEJ) of double strand breaks (DSBs). The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 system generates precise, locus-specific DSBs that can serve as substrates for HDR. We developed an in vivo visual reporter assay to quantify HDR-mediated events at single-cell resolution in zebrafish and used this system to identify small-molecule modulators that shift the DNA repair equilibrium in favor of HDR. By further optimizing the reaction environment and repair template, we achieved dramatic enhancement of HDR-mediated repair efficiency in zebrafish. Accordingly, under optimized conditions, inhibition of NHEJ with NU7441 enhanced HDR-mediated repair up to 13.4-fold. Importantly, we demonstrate that the increase in somatic HDR events correlates directly with germline transmission, permitting the efficient recovery of large seamlessly integrated DNA fragments in zebrafish.