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CRISPR/Cas9 genome editing in human hematopoietic stem cells

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CRISPR/Cas9 genome editing in human hematopoietic stem cells. / Bak, Rasmus; Dever, Daniel P; Porteus, Matthew H.

I: Nature Protocols (Print), Bind 13, Nr. 2, 2018, s. 358-376.

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

Harvard

Bak, R, Dever, DP & Porteus, MH 2018, 'CRISPR/Cas9 genome editing in human hematopoietic stem cells', Nature Protocols (Print), bind 13, nr. 2, s. 358-376. https://doi.org/10.1038/nprot.2017.143

APA

Bak, R., Dever, D. P., & Porteus, M. H. (2018). CRISPR/Cas9 genome editing in human hematopoietic stem cells. Nature Protocols (Print), 13(2), 358-376. https://doi.org/10.1038/nprot.2017.143

CBE

Bak R, Dever DP, Porteus MH. 2018. CRISPR/Cas9 genome editing in human hematopoietic stem cells. Nature Protocols (Print). 13(2):358-376. https://doi.org/10.1038/nprot.2017.143

MLA

Bak, Rasmus, Daniel P Dever og Matthew H Porteus. "CRISPR/Cas9 genome editing in human hematopoietic stem cells". Nature Protocols (Print). 2018, 13(2). 358-376. https://doi.org/10.1038/nprot.2017.143

Vancouver

Bak R, Dever DP, Porteus MH. CRISPR/Cas9 genome editing in human hematopoietic stem cells. Nature Protocols (Print). 2018;13(2):358-376. https://doi.org/10.1038/nprot.2017.143

Author

Bak, Rasmus ; Dever, Daniel P ; Porteus, Matthew H. / CRISPR/Cas9 genome editing in human hematopoietic stem cells. I: Nature Protocols (Print). 2018 ; Bind 13, Nr. 2. s. 358-376.

Bibtex

@article{e9fe616306454b668a0d42b29a393078,
title = "CRISPR/Cas9 genome editing in human hematopoietic stem cells",
abstract = "Genome editing via homologous recombination (HR) (gene targeting) in human hematopoietic stem cells (HSCs) has the power to reveal gene-function relationships and potentially transform curative hematological gene and cell therapies. However, there are no comprehensive and reproducible protocols for targeting HSCs for HR. Herein, we provide a detailed protocol for the production, enrichment, and in vitro and in vivo analyses of HR-targeted HSCs by combining CRISPR/Cas9 technology with the use of rAAV6 and flow cytometry. Using this protocol, researchers can introduce single-nucleotide changes into the genome or longer gene cassettes with the precision of genome editing. Along with our troubleshooting and optimization guidelines, researchers can use this protocol to streamline HSC genome editing at any locus of interest. The in vitro HSC-targeting protocol and analyses can be completed in 3 weeks, and the long-term in vivo HSC engraftment analyses in immunodeficient mice can be achieved in 16 weeks. This protocol enables manipulation of genes for investigation of gene functions during hematopoiesis, as well as for the correction of genetic mutations in HSC transplantation-based therapies for diseases such as sickle cell disease, β-thalassemia, and primary immunodeficiencies.",
keywords = "Journal Article",
author = "Rasmus Bak and Dever, {Daniel P} and Porteus, {Matthew H}",
year = "2018",
doi = "10.1038/nprot.2017.143",
language = "English",
volume = "13",
pages = "358--376",
journal = "Nature Protocols (Print)",
issn = "1754-2189",
publisher = "Nature Publishing Group",
number = "2",

}

RIS

TY - JOUR

T1 - CRISPR/Cas9 genome editing in human hematopoietic stem cells

AU - Bak, Rasmus

AU - Dever, Daniel P

AU - Porteus, Matthew H

PY - 2018

Y1 - 2018

N2 - Genome editing via homologous recombination (HR) (gene targeting) in human hematopoietic stem cells (HSCs) has the power to reveal gene-function relationships and potentially transform curative hematological gene and cell therapies. However, there are no comprehensive and reproducible protocols for targeting HSCs for HR. Herein, we provide a detailed protocol for the production, enrichment, and in vitro and in vivo analyses of HR-targeted HSCs by combining CRISPR/Cas9 technology with the use of rAAV6 and flow cytometry. Using this protocol, researchers can introduce single-nucleotide changes into the genome or longer gene cassettes with the precision of genome editing. Along with our troubleshooting and optimization guidelines, researchers can use this protocol to streamline HSC genome editing at any locus of interest. The in vitro HSC-targeting protocol and analyses can be completed in 3 weeks, and the long-term in vivo HSC engraftment analyses in immunodeficient mice can be achieved in 16 weeks. This protocol enables manipulation of genes for investigation of gene functions during hematopoiesis, as well as for the correction of genetic mutations in HSC transplantation-based therapies for diseases such as sickle cell disease, β-thalassemia, and primary immunodeficiencies.

AB - Genome editing via homologous recombination (HR) (gene targeting) in human hematopoietic stem cells (HSCs) has the power to reveal gene-function relationships and potentially transform curative hematological gene and cell therapies. However, there are no comprehensive and reproducible protocols for targeting HSCs for HR. Herein, we provide a detailed protocol for the production, enrichment, and in vitro and in vivo analyses of HR-targeted HSCs by combining CRISPR/Cas9 technology with the use of rAAV6 and flow cytometry. Using this protocol, researchers can introduce single-nucleotide changes into the genome or longer gene cassettes with the precision of genome editing. Along with our troubleshooting and optimization guidelines, researchers can use this protocol to streamline HSC genome editing at any locus of interest. The in vitro HSC-targeting protocol and analyses can be completed in 3 weeks, and the long-term in vivo HSC engraftment analyses in immunodeficient mice can be achieved in 16 weeks. This protocol enables manipulation of genes for investigation of gene functions during hematopoiesis, as well as for the correction of genetic mutations in HSC transplantation-based therapies for diseases such as sickle cell disease, β-thalassemia, and primary immunodeficiencies.

KW - Journal Article

UR - http://www.scopus.com/inward/record.url?scp=85041128425&partnerID=8YFLogxK

U2 - 10.1038/nprot.2017.143

DO - 10.1038/nprot.2017.143

M3 - Journal article

C2 - 29370156

VL - 13

SP - 358

EP - 376

JO - Nature Protocols (Print)

JF - Nature Protocols (Print)

SN - 1754-2189

IS - 2

ER -