Detection of Deleterious On-Target Effects after HDR-Mediated CRISPR Editing.

Autor:	Weisheit I; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany., Kroeger JA; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany., Malik R; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany., Klimmt J; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany., Crusius D; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany., Dannert A; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany., Dichgans M; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany; Munich Cluster for Systems Neurology (SyNergy), 81377 Munich, Germany., Paquet D; Institute for Stroke and Dementia Research (ISD), University Hospital, LMU Munich, 81377 Munich, Germany; Graduate School of Systemic Neurosciences, LMU Munich, 82152 Planegg-Martinsried, Germany; Munich Cluster for Systems Neurology (SyNergy), 81377 Munich, Germany. Electronic address: dominik.paquet@med.uni-muenchen.de.
Jazyk:	angličtina
Zdroj:	Cell reports [Cell Rep] 2020 May 26; Vol. 31 (8), pp. 107689.
DOI:	10.1016/j.celrep.2020.107689
Abstrakt:	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. Competing Interests: Declaration of Interests The authors declare no competing interests. (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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