Efficient Ablation of Genes in Human Hematopoietic Stem and Effector Cells using CRISPR/Cas9.

Pankaj K Mandal, Leonardo M R Ferreira, Ryan Collins, Torsten B Meissner, Christian L Boutwell, Max Friesen, Vladimir Vrbanac, Brian S Garrison, Alexei Stortchevoi, David Bryder, Kiran Musunuru, Harrison Brand, Andrew M Tager, Todd M Allen, Michael E Talkowski, Derrick J Rossi, Chad A Cowan

Research output: Contribution to journalArticlepeer-review

293 Citations (SciVal)


Genome editing via CRISPR/Cas9 has rapidly become the tool of choice by virtue of its efficacy and ease of use. However, CRISPR/Cas9-mediated genome editing in clinically relevant human somatic cells remains untested. Here, we report CRISPR/Cas9 targeting of two clinically relevant genes, B2M and CCR5, in primary human CD4(+) T cells and CD34(+) hematopoietic stem and progenitor cells (HSPCs). Use of single RNA guides led to highly efficient mutagenesis in HSPCs but not in T cells. A dual guide approach improved gene deletion efficacy in both cell types. HSPCs that had undergone genome editing with CRISPR/Cas9 retained multilineage potential. We examined predicted on- and off-target mutations via target capture sequencing in HSPCs and observed low levels of off-target mutagenesis at only one site. These results demonstrate that CRISPR/Cas9 can efficiently ablate genes in HSPCs with minimal off-target mutagenesis, which could have broad applicability for hematopoietic cell-based therapy.
Original languageEnglish
Pages (from-to)643-652
JournalCell Stem Cell
Issue number5
Publication statusPublished - 2014

Bibliographical note

The information about affiliations in this record was updated in December 2015.
The record was previously connected to the following departments: Stem Cell Aging (013212073)

Subject classification (UKÄ)

  • Cell Biology


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