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Generation and characterization of a human iPSC cell line expressing inducible Cas9 in the “safe harbor” AAVS1 locus
oleh: Julio Castaño, Clara Bueno, Senda Jiménez-Delgado, Heleia Roca-Ho, Mario F. Fraga, Agustín F. Fernandez, Mahito Nakanishi, Raúl Torres-Ruiz, Sandra Rodríguez-Perales, Pablo Menéndez
Format: | Article |
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Diterbitkan: | Elsevier 2017-05-01 |
Deskripsi
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.