Evaluating the function of murine quiescent hematopoietic stem cells following non-homologous end joining-based genome editing
Related Experiment Video
Updated: Jul 27, 2025

CRISPR/Cas9 Gene Editing of Hematopoietic Stem and Progenitor Cells for Gene Therapy Applications
Published on: August 9, 2022
Kohei Shiroshita1, Hiroshi Kobayashi1, Keiyo Takubo1
1Department of Stem Cell Biology, Research Institute, National Center for Global Health and Medicine, Tokyo 162-8685, Japan.
Abstract:
Preculture is indispensable for achieving highly efficient non-homologous end joining (NHEJ)-based genome editing. Here, we present a protocol for optimizing genome editing conditions for murine hematopoietic stem cells (HSCs) and evaluating their function following NHEJ-based genome editing. We describe steps for sgRNA preparation, cell sorting, preculture, and electroporation. We then detail post-editing culture and transplanting of bone marrow. This protocol can be used to study genes related to HSC quiescence. For complete details on the use and execution of this protocol, please refer to Shiroshita et al.1.
Related Concept Videos
Multipotency of Hematopoietic Stem Cells
Regulation of Hematopoietic Stem Cells
Homologous Recombination
In-vitro Mutagenesis

