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CRISPR/Cas-mediated knockin in human pluripotent stem cells

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Abstract

Fluorescent reporter and epitope-tagged human pluripotent stem cells (hPSCs) greatly facilitate studies on the pluripotency and differentiation characteristics of these cells. Unfortunately traditional procedures to generate such lines are hampered by a low targeting effi ciency that necessitates a lengthy process of selection followed by the removal of the selection cassette. Here we describe a procedure to generate fl uorescent reporter and epitope tagged hPSCs in an effi cient one-step process using the CRISPR/Cas technology. Although the method described uses our recently developed iCRISPR platform, the protocols can be adapted for general use with CRISPR/Cas or other engineered nucleases. The transfection procedures described could also be used for additional applications, such as overexpression or lineage tracing studies.

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Verma, N., Zhu, Z., & Huangfu, D. (2017). CRISPR/Cas-mediated knockin in human pluripotent stem cells. In Methods in Molecular Biology (Vol. 1513, pp. 119–140). Humana Press Inc. https://doi.org/10.1007/978-1-4939-6539-7_9

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