RRC ID 62885
Author Maurissen TL, Woltjen K.
Title Synergistic gene editing in human iPS cells via cell cycle and DNA repair modulation.
Journal Nat Commun
Abstract Precise gene editing aims at generating single-nucleotide modifications to correct or model human disease. However, precision editing with nucleases such as CRIPSR-Cas9 has seen limited success due to poor efficiency and limited practicality. Here, we establish a fluorescent DNA repair assay in human induced pluripotent stem (iPS) cells to visualize and quantify the frequency of DNA repair outcomes during monoallelic and biallelic targeting. We found that modulating both DNA repair and cell cycle phase via defined culture conditions and small molecules synergistically enhanced the frequency of homology-directed repair (HDR). Notably, targeting in homozygous reporter cells results in high levels of editing with a vast majority of biallelic HDR outcomes. We then leverage efficient biallelic HDR with mixed ssODN repair templates to generate heterozygous mutations. Synergistic gene editing represents an effective strategy to generate precise genetic modifications in human iPS cells.
Volume 11(1)
Pages 2876
Published 2020-6-8
DOI 10.1038/s41467-020-16643-5
PII 10.1038/s41467-020-16643-5
PMID 32513994
PMC PMC7280248
MeSH Alleles Amino Acid Sequence Base Sequence Cell Cycle* Cold-Shock Response DNA Repair* Fluorescence Gene Editing* Genetic Loci Green Fluorescent Proteins / chemistry Green Fluorescent Proteins / metabolism Homozygote Humans Induced Pluripotent Stem Cells / cytology* Induced Pluripotent Stem Cells / metabolism* Mutation / genetics
IF 12.121
Human and Animal Cells 409B2(HPS0076)