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Lookup NU author(s): Sara Cuevas Ocaña, Dr George Schlossmacher, Professor Neil PerkinsORCiD, Dr Michael Gray
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© 2023 by the authors. Introducing or correcting disease-causing mutations through genome editing in human pluripotent stem cells (hPSCs) followed by tissue-specific differentiation provide sustainable models of multiorgan diseases, such as cystic fibrosis (CF). However, low editing efficiency resulting in extended cell culture periods and the use of specialised equipment for fluorescence activated cell sorting (FACS) make hPSC genome editing still challenging. We aimed to investigate whether a combination of cell cycle synchronisation, single-stranded oligodeoxyribonucleotides, transient selection, manual clonal isolation, and rapid screening can improve the generation of correctly modified hPSCs. Here, we introduced the most common CF mutation, ΔF508, into the CFTR gene, using TALENs into hPSCs, and corrected the W1282X mutation using CRISPR-Cas9, in human-induced PSCs. This relatively simple method achieved up to 10% efficiency without the need for FACS, generating heterozygous and homozygous gene edited hPSCs within 3–6 weeks in order to understand genetic determinants of disease and precision medicine.
Author(s): Cuevas-Ocana S, Yang JY, Aushev M, Schlossmacher G, Bear CE, Hannan NRF, Perkins ND, Rossant J, Wong AP, Gray MA
Publication type: Article
Publication status: Published
Journal: International Journal of Molecular Sciences
Year: 2023
Volume: 24
Issue: 12
Online publication date: 17/06/2023
Acceptance date: 09/06/2023
ISSN (print): 1661-6596
ISSN (electronic): 1422-0067
Publisher: MDPI
URL: https://doi.org/10.3390/ijms241210266
DOI: 10.3390/ijms241210266
PubMed id: 37373413
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