Detection of Deleterious On-Target Effects after HDR-Mediated CRISPR Editing.
APP
CRISPR
HDR
genome editing
homology-directed repair
iPSCs
induced pluripotent stem cells
loss-of-heterozygosity
on-target effects
qPCR
Journal
Cell reports
ISSN: 2211-1247
Titre abrégé: Cell Rep
Pays: United States
ID NLM: 101573691
Informations de publication
Date de publication:
26 05 2020
26 05 2020
Historique:
received:
27
11
2019
revised:
22
04
2020
accepted:
04
05
2020
entrez:
28
5
2020
pubmed:
28
5
2020
medline:
22
5
2021
Statut:
ppublish
Résumé
CRISPR genome editing is a promising tool for translational research but can cause undesired editing outcomes, both on target at the edited locus and off target at other genomic loci. Here, we investigate the occurrence of deleterious on-target effects (OnTEs) in human stem cells after insertion of disease-related mutations by homology-directed repair (HDR) and gene editing using non-homologous end joining (NHEJ). We identify large, mono-allelic genomic deletions and loss-of-heterozygosity escaping standard quality controls in up to 40% of edited clones. To reliably detect such events, we describe simple, low-cost, and broadly applicable quantitative genotyping PCR (qgPCR) and single-nucleotide polymorphism (SNP) genotyping-based tools and suggest their usage as additional quality controls after editing. This will help to ensure the integrity of edited loci and increase the reliability of CRISPR editing.
Identifiants
pubmed: 32460021
pii: S2211-1247(20)30642-2
doi: 10.1016/j.celrep.2020.107689
pii:
doi:
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
107689Informations de copyright
Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Declaration of Interests The authors declare no competing interests.