Protocol for CRISPR-Cas12a genome editing of protein tyrosine phosphatases in human pluripotent stem cells and functional β-like cell generation
Gene editing of human pluripotent stem cells is a promising approach for developing targeted gene therapies for metabolic diseases. Here, we present a protocol for generating a CRISPR-Cas12a gene knockout of protein tyrosine phosphatases in human embryonic stem cells. We describe steps for differentiating the edited clones into pancreatic islet-like spheroids rich in β-like cells. We then detail procedures for implanting these spheroids under the murine kidney capsule for in vivo maturation.
Authors
- Mariana Nunes (ORCID: https://orcid.org/0000-0002-4733-5778)
- André Dias (ORCID: https://orcid.org/0000-0003-3337-6373)
- Stephanie Talamantes (ORCID: https://orcid.org/0000-0001-8211-4834)
- Esteban N. Gurzov (ORCID: https://orcid.org/0000-0003-4642-0273)
- Valerie Vandenbempt (ORCID: https://orcid.org/0000-0003-3104-8499)
- Mayank Bansal (ORCID: https://orcid.org/0000-0002-5798-5352)
- Javier Negueruela (ORCID: https://orcid.org/0000-0001-7967-5067)
- Francisco Ribeiro-Costa
Institutions
- Université Libre de Bruxelles (BE)
Publication Details
- Journal
- STAR Protocols
- Published
- 2024-09-01
- DOI
- https://doi.org/10.1016/j.xpro.2024.103297
- Citations
- 4
- Primary Topic
- Pancreatic function and diabetes
- Type
- article
- Field-Weighted Citation Impact
- 2.29
Funders
- U.S. Department of Agriculture
- Helsingin Yliopisto
- Fonds De La Recherche Scientifique - FNRS
- H2020 European Research Council