TitleGenome Engineering of Hematopoietic Stem Cells Using CRISPR/Cas9 System.
Publication TypeJournal Article
Year of Publication2022
AuthorsDevaraju N, Rajendiran V, Ravi NSam, Mohankumar KM
JournalMethods Mol Biol
Date Published2022
KeywordsAnimals, CRISPR-Cas Systems, Gene Editing, Hematopoietic Stem Cell Transplantation, Hematopoietic Stem Cells, Mice, Transplantation, Autologous

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.

Alternate JournalMethods Mol Biol
PubMed ID35507170