期刊
CELL STEM CELL
卷 15, 期 5, 页码 643-652出版社
CELL PRESS
DOI: 10.1016/j.stem.2014.10.004
关键词
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资金
- Portuguese Foundation for Science and Technology
- National Institutes of Health [P01-AI104715, MH095867, R01HL107630, U01DK072473, R01DK097768, U01HL100408]
- Simons Foundation
- March of Dimes
- Leona M. and Harry B. Helmsley Charitable Trust
- Jane Brock-Wilson Fund
- Harvard Stem Cell Institute
- Harvard University
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.
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