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Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells
Mutations that accumulate in self-renewing hematopoietic stem and progenitor cells (HSPCs) can cause severe blood disorders. To model such disorders in mice, we developed a CRISPR/Cas9/adeno-associated virus (AAV)-based system to knock in and repair genes by homologous recombination in mouse HSPCs....
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7357675/ https://www.ncbi.nlm.nih.gov/pubmed/32685932 http://dx.doi.org/10.1016/j.xpro.2020.100028 |
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author | Tran, Ngoc Tung Trombke, Janine Rajewsky, Klaus Chu, Van Trung |
author_facet | Tran, Ngoc Tung Trombke, Janine Rajewsky, Klaus Chu, Van Trung |
author_sort | Tran, Ngoc Tung |
collection | PubMed |
description | Mutations that accumulate in self-renewing hematopoietic stem and progenitor cells (HSPCs) can cause severe blood disorders. To model such disorders in mice, we developed a CRISPR/Cas9/adeno-associated virus (AAV)-based system to knock in and repair genes by homologous recombination in mouse HSPCs. Here, we provide a step-by-step protocol to achieve high efficiency of gene knockin in mouse HSPCs, while maintaining engraftment capacity. This approach enables the functional study of hematopoietic disease mutations in vivo, without requiring germline mutagenesis. For complete details on the use and execution of this protocol, please refer to Tran et al. (2019). |
format | Online Article Text |
id | pubmed-7357675 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-73576752020-07-17 Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells Tran, Ngoc Tung Trombke, Janine Rajewsky, Klaus Chu, Van Trung STAR Protoc Protocol Mutations that accumulate in self-renewing hematopoietic stem and progenitor cells (HSPCs) can cause severe blood disorders. To model such disorders in mice, we developed a CRISPR/Cas9/adeno-associated virus (AAV)-based system to knock in and repair genes by homologous recombination in mouse HSPCs. Here, we provide a step-by-step protocol to achieve high efficiency of gene knockin in mouse HSPCs, while maintaining engraftment capacity. This approach enables the functional study of hematopoietic disease mutations in vivo, without requiring germline mutagenesis. For complete details on the use and execution of this protocol, please refer to Tran et al. (2019). Elsevier 2020-06-03 /pmc/articles/PMC7357675/ /pubmed/32685932 http://dx.doi.org/10.1016/j.xpro.2020.100028 Text en © 2020 The Authors http://creativecommons.org/licenses/by/4.0/ This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Protocol Tran, Ngoc Tung Trombke, Janine Rajewsky, Klaus Chu, Van Trung Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title | Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title_full | Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title_fullStr | Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title_full_unstemmed | Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title_short | Protocol for Efficient CRISPR/Cas9/AAV-Mediated Homologous Recombination in Mouse Hematopoietic Stem and Progenitor Cells |
title_sort | protocol for efficient crispr/cas9/aav-mediated homologous recombination in mouse hematopoietic stem and progenitor cells |
topic | Protocol |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7357675/ https://www.ncbi.nlm.nih.gov/pubmed/32685932 http://dx.doi.org/10.1016/j.xpro.2020.100028 |
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