Efficient Precision Genome Editing in iPSCs via Genetic Co-targeting with Selection

Genome editing in induced pluripotent stem cells is currently hampered by the laborious and expensive nature of identifying homology-directed repair (HDR)-modified cells. We present an approach where isolation of cells bearing a selectable, HDR-mediated editing event at one locus enriches for HDR-me...

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Published in:Stem cell reports 2017-03, Vol.8 (3), p.491-499
Main Authors: Mitzelfelt, Katie A., McDermott-Roe, Chris, Grzybowski, Michael N., Marquez, Maribel, Kuo, Chieh-Ti, Riedel, Michael, Lai, Shuping, Choi, Melinda J., Kolander, Kurt D., Helbling, Daniel, Dimmock, David P., Battle, Michele A., Jou, Chuanchau J., Tristani-Firouzi, Martin, Verbsky, James W., Benjamin, Ivor J., Geurts, Aron M.
Format: Article
Language:English
Subjects:
Cell Line
CRISPR-Cas Systems
DNA End-Joining Repair
Gene Editing
Gene Knock-In Techniques
Gene Targeting
Genetic Vectors
Genome, Human
High-Throughput Nucleotide Sequencing
Humans
Induced Pluripotent Stem Cells - cytology
Induced Pluripotent Stem Cells - drug effects
Induced Pluripotent Stem Cells - metabolism
Recombinational DNA Repair
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Summary:Genome editing in induced pluripotent stem cells is currently hampered by the laborious and expensive nature of identifying homology-directed repair (HDR)-modified cells. We present an approach where isolation of cells bearing a selectable, HDR-mediated editing event at one locus enriches for HDR-mediated edits at additional loci. This strategy, called co-targeting with selection, improves the probability of isolating cells bearing HDR-mediated variants and accelerates the production of disease models. [Display omitted] •Increases the efficiency of genome editing in human iPSCs•Enhances detectability of variants of interest derived by homology-directed repair•Is a simple, scalable, and adaptable strategy for knocking in variants of interest The potential of CRISPR/Cas9-based genome editing in iPSCs is currently hampered by the laborious nature of identifying cells modified by the lesser-used homology-directed repair (HDR) pathway. Geurts, McDermott-Roe, and colleagues present a straightforward approach, co-targeting with selection, based on co-modification that enriches for cells undergoing HDR thereby improving targeting efficiencies.
ISSN:2213-6711
2213-6711
DOI:10.1016/j.stemcr.2017.01.021