Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells

We present here a protocol for biallelic tagging of an endogenous gene in human cells using CRISPR-Cas9 editing technology. Using RIF1 as an example, we describe tagging the gene with a mini-auxin-inducible degron and a green fluorescent protein at its C terminus. We detail steps for preparing and d...

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Detalles Bibliográficos
Autores principales: Kong, Nannan, Chan, Ying Wai
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2023
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10241970/
https://www.ncbi.nlm.nih.gov/pubmed/37252842
http://dx.doi.org/10.1016/j.xpro.2023.102286
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author Kong, Nannan
Chan, Ying Wai
author_facet Kong, Nannan
Chan, Ying Wai
author_sort Kong, Nannan
collection PubMed
description We present here a protocol for biallelic tagging of an endogenous gene in human cells using CRISPR-Cas9 editing technology. Using RIF1 as an example, we describe tagging the gene with a mini-auxin-inducible degron and a green fluorescent protein at its C terminus. We detail steps for preparing and designing the sgRNA and homologous repair template, and clone selection and verification. For complete details on the use and execution of this protocol, please refer to Kong et al.(1)
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spelling pubmed-102419702023-06-07 Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells Kong, Nannan Chan, Ying Wai STAR Protoc Protocol We present here a protocol for biallelic tagging of an endogenous gene in human cells using CRISPR-Cas9 editing technology. Using RIF1 as an example, we describe tagging the gene with a mini-auxin-inducible degron and a green fluorescent protein at its C terminus. We detail steps for preparing and designing the sgRNA and homologous repair template, and clone selection and verification. For complete details on the use and execution of this protocol, please refer to Kong et al.(1) Elsevier 2023-05-29 /pmc/articles/PMC10241970/ /pubmed/37252842 http://dx.doi.org/10.1016/j.xpro.2023.102286 Text en © 2023 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Kong, Nannan
Chan, Ying Wai
Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title_full Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title_fullStr Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title_full_unstemmed Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title_short Protocol for biallelic tagging of an endogenous gene using CRISPR-Cas9 in human cells
title_sort protocol for biallelic tagging of an endogenous gene using crispr-cas9 in human cells
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10241970/
https://www.ncbi.nlm.nih.gov/pubmed/37252842
http://dx.doi.org/10.1016/j.xpro.2023.102286
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