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Stem Cell Res. 2017 May;21:137-140. doi: 10.1016/j.scr.2017.04.011. Epub 2017 Apr 22.

Generation and characterization of a human iPSC cell line expressing inducible Cas9 in the "safe harbor" AAVS1 locus.

Author information

1
Josep Carreras Leukemia Research Institute, Department of Biomedicine, School of Medicine, University of Barcelona, Barcelona, Spain; Center for Networked Biomedical Research on Cancer (CIBERONC), ISCIII, Madrid, Spain. Electronic address: jcastano@carrerasresearch.org.
2
Josep Carreras Leukemia Research Institute, Department of Biomedicine, School of Medicine, University of Barcelona, Barcelona, Spain; Center for Networked Biomedical Research on Cancer (CIBERONC), ISCIII, Madrid, Spain.
3
Center of Regenerative Medicine in Barcelona, Barcelona Biomedical Research Park, Barcelona, Spain; Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBERBBN), ISCIII, Madrid, Spain.
4
Nanomaterials and Nanotechnology Research Center (CINN-CSIC), Universidad de Oviedo-Principado de Asturias, Spain.
5
Cancer Epigenetics Laboratory, Institute of Oncology of Asturias (IUOPA), Hospital Universitario Central de Asturias HUCA-FINBA, Universidad de Oviedo, Spain.
6
National Institute of Advanced Industrial Science and Technology, Tsukuba, Ibaraki, Japan.
7
Josep Carreras Leukemia Research Institute, Department of Biomedicine, School of Medicine, University of Barcelona, Barcelona, Spain.
8
Cytogenetics Group, Centro Nacional de Investigaciones Oncológicas (CNIO), Madrid, Spain.
9
Josep Carreras Leukemia Research Institute, Department of Biomedicine, School of Medicine, University of Barcelona, Barcelona, Spain; Center for Networked Biomedical Research on Cancer (CIBERONC), ISCIII, Madrid, Spain; Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain.

Abstract

We report the generation-characterization of a fetal liver (FL) B-cell progenitor (BCP)-derived human induced pluripotent stem cell (hiPSC) line CRISPR/Cas9-edited to carry/express a single copy of doxycycline-inducible Cas9 gene in the "safe locus" AAVS1 (iCas9-FL-BCP-hiPSC). Gene-edited iPSCs remained pluripotent after CRISPR/Cas9 genome-edition. Correct genomic integration of a unique copy of Cas9 was confirmed by PCR and Southern blot. Cas9 was robustly and specifically expressed on doxycycline exposure. T7-endonuclease assay demonstrated that iCas9 induces robust gene-edition when gRNAs against hematopoietic transcription factors were tested. This iCas9-FL-BCP-hiPSC will facilitate gene-editing approaches for studies on developmental biology, drug screening and disease modeling.

PMID:
28677529
PMCID:
PMC5446316
DOI:
10.1016/j.scr.2017.04.011
[Indexed for MEDLINE]
Free PMC Article

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