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Genome Engineering of Hematopoietic Stem Cells Using CRISPR/Cas9 System

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Stem Cell Assays

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2429))

Abstract

Ex vivo genetic manipulation of autologous hematopoietic stem and progenitor cells (HSPCs) is a viable strategy for the treatment of hematologic and primary immune disorders. Targeted genome editing of HSPCs using the CRISPR-Cas9 system provides an effective platform to edit the desired genomic locus for therapeutic purposes with minimal off-target effects. In this chapter, we describe the detailed methodology for the CRISPR-Cas9 mediated gene knockout, deletion, addition, and correction in human HSPCs by viral and nonviral approaches. We also present a comprehensive protocol for the analysis of genome modified HSPCs toward the erythroid and megakaryocyte lineage in vitro and the long-term multilineage reconstitution capacity in the recently developed NBSGW mouse model that supports human erythropoiesis.

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Authors and Affiliations

  1. Centre for Stem Cell Research (a unit of inStem, Bangalore), Christian Medical College Campus, Bagayam, Vellore, Tamil Nadu, India

    Nivedhitha Devaraju, Vignesh Rajendiran, Nithin Sam Ravi & Kumarasamypet M. Mohankumar

  2. Manipal Academy of Higher Education, Mangalore, Karnataka, India

    Nivedhitha Devaraju

Authors
  1. Nivedhitha Devaraju
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  2. Vignesh Rajendiran
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  3. Nithin Sam Ravi
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  4. Kumarasamypet M. Mohankumar
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Corresponding author

Correspondence to Kumarasamypet M. Mohankumar .

Editor information

Editors and Affiliations

  1. Stem Cell and Cancer Biology Laboratory, Division of Experimental Medicine and Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA

    Nagarajan Kannan

  2. Hull York Medical School, York, UK

    Philip Beer

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Devaraju, N., Rajendiran, V., Ravi, N.S., Mohankumar, K.M. (2022). Genome Engineering of Hematopoietic Stem Cells Using CRISPR/Cas9 System. In: Kannan, N., Beer, P. (eds) Stem Cell Assays. Methods in Molecular Biology, vol 2429. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1979-7_20

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  • DOI: https://doi.org/10.1007/978-1-0716-1979-7_20

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1978-0

  • Online ISBN: 978-1-0716-1979-7

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