CRISPR/Cas9-Mediated Genome Editing to Generate Clonal iPSC Lines

The ability to reprogram somatic cells into induced pluripotent stem cells (iPSCs) was developed in 2006 and represented a major breakthrough in stem cell research. A more recent milestone in biomedical research was reached in 2013 when the CRISPR/Cas9 system was used to edit the genome of mammalian...

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Published inMethods in molecular biology (Clifton, N.J.)
Main Authors Sanjurjo-Soriano, Carla, Erkilic, Nejla, Mamaeva, Daria, Kalatzis, Vasiliki
Format Journal Article
LanguageEnglish
Published United States 24.03.2021
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Abstract The ability to reprogram somatic cells into induced pluripotent stem cells (iPSCs) was developed in 2006 and represented a major breakthrough in stem cell research. A more recent milestone in biomedical research was reached in 2013 when the CRISPR/Cas9 system was used to edit the genome of mammalian cells. The coupling of both human (h)iPSCs and CRISPR/Cas9 technology offers great promise for cell therapy and regenerative medicine. However, several limitations including time and labor consumption, efficiency and efficacy of the system, and the potential off-targets effects induced by the Cas9 nuclease still need to be addressed. Here, we describe a detailed method for easily engineering genetic changes in hiPSCs, using a nucleofection-mediated protocol to deliver the CRISPR/Cas9 components into the cells, and discuss key points to be considered when designing your experiment. The clonal, genome-edited hiPSC line generated via our method can be directly used for downstream applications.
AbstractList The ability to reprogram somatic cells into induced pluripotent stem cells (iPSCs) was developed in 2006 and represented a major breakthrough in stem cell research. A more recent milestone in biomedical research was reached in 2013 when the CRISPR/Cas9 system was used to edit the genome of mammalian cells. The coupling of both human (h)iPSCs and CRISPR/Cas9 technology offers great promise for cell therapy and regenerative medicine. However, several limitations including time and labor consumption, efficiency and efficacy of the system, and the potential off-targets effects induced by the Cas9 nuclease still need to be addressed. Here, we describe a detailed method for easily engineering genetic changes in hiPSCs, using a nucleofection-mediated protocol to deliver the CRISPR/Cas9 components into the cells, and discuss key points to be considered when designing your experiment. The clonal, genome-edited hiPSC line generated via our method can be directly used for downstream applications.
Author Sanjurjo-Soriano, Carla
Kalatzis, Vasiliki
Erkilic, Nejla
Mamaeva, Daria
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  givenname: Nejla
  surname: Erkilic
  fullname: Erkilic, Nejla
  organization: University of Montpellier, Montpellier, France
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  givenname: Daria
  surname: Mamaeva
  fullname: Mamaeva, Daria
  organization: University of Montpellier, Montpellier, France
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  givenname: Vasiliki
  surname: Kalatzis
  fullname: Kalatzis, Vasiliki
  email: vasiliki.kalatzis@inserm.fr, vasiliki.kalatzis@inserm.fr
  organization: University of Montpellier, Montpellier, France. vasiliki.kalatzis@inserm.fr
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Keywords Single-cell sorting
iPSCs
Genome editing
CRISPR/Cas9
Nucleofection
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Snippet The ability to reprogram somatic cells into induced pluripotent stem cells (iPSCs) was developed in 2006 and represented a major breakthrough in stem cell...
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Title CRISPR/Cas9-Mediated Genome Editing to Generate Clonal iPSC Lines
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