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Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts

Human induced pluripotent stem cells (hiPSCs) allow in vitro study of genetic diseases and hold potential for personalized stem cell therapy. Gene editing, precisely modifying specifically targeted loci, represents a valuable tool for different hiPSC applications. This is especially useful in monoge...

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Autores principales: Jalil, Sami, Keskinen, Timo, Maldonado, Rocío, Sokka, Joonas, Trokovic, Ras, Otonkoski, Timo, Wartiovaara, Kirmo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8693657/
https://www.ncbi.nlm.nih.gov/pubmed/34822772
http://dx.doi.org/10.1016/j.stemcr.2021.10.017
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author Jalil, Sami
Keskinen, Timo
Maldonado, Rocío
Sokka, Joonas
Trokovic, Ras
Otonkoski, Timo
Wartiovaara, Kirmo
author_facet Jalil, Sami
Keskinen, Timo
Maldonado, Rocío
Sokka, Joonas
Trokovic, Ras
Otonkoski, Timo
Wartiovaara, Kirmo
author_sort Jalil, Sami
collection PubMed
description Human induced pluripotent stem cells (hiPSCs) allow in vitro study of genetic diseases and hold potential for personalized stem cell therapy. Gene editing, precisely modifying specifically targeted loci, represents a valuable tool for different hiPSC applications. This is especially useful in monogenic diseases to dissect the function of unknown mutations or to create genetically corrected, patient-derived hiPSCs. Here we describe a highly efficient method for simultaneous base editing and reprogramming of fibroblasts employing a CRISPR-Cas9 adenine base editor. As a proof of concept, we apply this approach to generate gene-edited hiPSCs from skin biopsies of four patients carrying a Finnish-founder pathogenic point mutation in either NOTCH3 or LDLR genes. We also show LDLR activity restoration after the gene correction. Overall, this method yields tens of gene-edited hiPSC monoclonal lines with unprecedented efficiency and robustness while considerably reducing the cell culture time and thus the risk for in vitro alterations.
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spelling pubmed-86936572022-01-04 Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts Jalil, Sami Keskinen, Timo Maldonado, Rocío Sokka, Joonas Trokovic, Ras Otonkoski, Timo Wartiovaara, Kirmo Stem Cell Reports Resource Human induced pluripotent stem cells (hiPSCs) allow in vitro study of genetic diseases and hold potential for personalized stem cell therapy. Gene editing, precisely modifying specifically targeted loci, represents a valuable tool for different hiPSC applications. This is especially useful in monogenic diseases to dissect the function of unknown mutations or to create genetically corrected, patient-derived hiPSCs. Here we describe a highly efficient method for simultaneous base editing and reprogramming of fibroblasts employing a CRISPR-Cas9 adenine base editor. As a proof of concept, we apply this approach to generate gene-edited hiPSCs from skin biopsies of four patients carrying a Finnish-founder pathogenic point mutation in either NOTCH3 or LDLR genes. We also show LDLR activity restoration after the gene correction. Overall, this method yields tens of gene-edited hiPSC monoclonal lines with unprecedented efficiency and robustness while considerably reducing the cell culture time and thus the risk for in vitro alterations. Elsevier 2021-11-24 /pmc/articles/PMC8693657/ /pubmed/34822772 http://dx.doi.org/10.1016/j.stemcr.2021.10.017 Text en © 2021 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Resource
Jalil, Sami
Keskinen, Timo
Maldonado, Rocío
Sokka, Joonas
Trokovic, Ras
Otonkoski, Timo
Wartiovaara, Kirmo
Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title_full Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title_fullStr Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title_full_unstemmed Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title_short Simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
title_sort simultaneous high-efficiency base editing and reprogramming of patient fibroblasts
topic Resource
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8693657/
https://www.ncbi.nlm.nih.gov/pubmed/34822772
http://dx.doi.org/10.1016/j.stemcr.2021.10.017
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