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Genome Editing in Human Pluripotent Stem Cells Using Site-Specific Nucleases

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Chromosomal Mutagenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1239))

Abstract

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) (Thomson, Science 282:1145–1147, 1998; Takahashi et al. Cell 131:861–872, 2007), collectively referred to as pluripotent stem cells (hPSCs), are currently used in disease modeling to address questions specific to humans and to complement our insight gained from model organisms (Soldner et al. Cell 146:318–331, 2011; Soldner and Jaenisch, Science 338:1155–1156, 2012). Recently, genetic engineering using site-specific nucleases has been established in hPSCs (Hockemeyer et al. Nat Biotechnol 27:851–857, 2009; Hockemeyer et al., Nat Biotechnol 29:731–734, 2011; Zou et al., Cell Stem Cell 5:97–110, 2011; Yusa et al., Nature 478:391–394, 2011; DeKelver et al., Genome Res 20:1133–1142, 2010), allowing a level of genetic control previously limited to model systems. Thus, we can now perform targeted gene knockouts, generate tissue-specific cell lineage reporters, overexpress genes from a defined locus, and introduce and repair single point mutations in hPSCs. This ability to genetically engineer pluripotent stem cells will significantly facilitate the study of human disease in a defined genetic context. Here we outline protocols for efficient gene targeting in hPSCs.

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Author information

Authors and Affiliations

  1. Department of Molecular and Cell Biology, University of California, Berkeley, 400B Li Ka Shing Center, Berkeley, CA, 94720-3370, USA

    Kunitoshi Chiba & Dirk Hockemeyer

Authors
  1. Kunitoshi Chiba
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  2. Dirk Hockemeyer
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Corresponding author

Correspondence to Dirk Hockemeyer .

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Editors and Affiliations

  1. Department of Genetics, Genome Engineering and iPSC Center, Washington University School of Medicine, St. Louis, Missouri, USA

    Shondra M. Pruett-Miller

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Chiba, K., Hockemeyer, D. (2015). Genome Editing in Human Pluripotent Stem Cells Using Site-Specific Nucleases. In: Pruett-Miller, S. (eds) Chromosomal Mutagenesis. Methods in Molecular Biology, vol 1239. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1862-1_15

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  • DOI: https://doi.org/10.1007/978-1-4939-1862-1_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1861-4

  • Online ISBN: 978-1-4939-1862-1

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