In this chapter, we present an optimized CRISPR/Cas9 RNP nucleofection approach for gene knockout (KO) in hematopoietic stem and progenitor cells (HSPCs). With experimentally proved active locus-specific sgRNAs, we routinely reach over 80% gene KO in HSPCs, thus avoiding the need for cell sorting or enrichment of targeted cell population. Additionally, we provide a protocol for in vitro granulocytic differentiation of HSPCs after gene KO and detailed description of granulocyte function tests which can be applied to study the effects of a particular gene KO.
CITATION STYLE
Mir, P., Ritter, M., Welte, K., Skokowa, J., & Klimiankou, M. (2020). Gene Knockout in Hematopoietic Stem and Progenitor Cells Followed by Granulocytic Differentiation. In Methods in Molecular Biology (Vol. 2115, pp. 455–469). Humana Press Inc. https://doi.org/10.1007/978-1-0716-0290-4_26
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