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BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B
BACKGROUND: Base Editing is a precise genome editing method that uses a deaminase-Cas9 fusion protein to mutate cytidine to thymidine in target DNA in situ without the generation of a double-strand break. However, the efficient enrichment of genetically modified cells using this technique is limited...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6309101/ https://www.ncbi.nlm.nih.gov/pubmed/30593278 http://dx.doi.org/10.1186/s12915-018-0617-1 |
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author | Coelho, Matthew A. Li, Songyuan Pane, Luna Simona Firth, Mike Ciotta, Giovanni Wrigley, Jonathan D. Cuomo, Maria Emanuela Maresca, Marcello Taylor, Benjamin J. M. |
author_facet | Coelho, Matthew A. Li, Songyuan Pane, Luna Simona Firth, Mike Ciotta, Giovanni Wrigley, Jonathan D. Cuomo, Maria Emanuela Maresca, Marcello Taylor, Benjamin J. M. |
author_sort | Coelho, Matthew A. |
collection | PubMed |
description | BACKGROUND: Base Editing is a precise genome editing method that uses a deaminase-Cas9 fusion protein to mutate cytidine to thymidine in target DNA in situ without the generation of a double-strand break. However, the efficient enrichment of genetically modified cells using this technique is limited by the ability to detect such events. RESULTS: We have developed a Base Editing FLuorescent Activity REporter (BE-FLARE), which allows for the enrichment of cells that have undergone editing of target loci based on a fluorescence shift from BFP to GFP. We used BE-FLARE to evaluate the editing efficiency of APOBEC3A and APOBEC3B family members as alternatives deaminase domains to the rat APOBEC1 domain used in base editor 3 (BE3). We identified human APOBEC3A and APOBEC3B as highly efficient cytidine deaminases for base editing applications with unique properties. CONCLUSIONS: Using BE-FLARE to report on the efficiency and precision of editing events, we outline workflows for the accelerated generation of genetically engineered cell models and the discovery of alternative base editors. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12915-018-0617-1) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-6309101 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-63091012019-01-03 BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B Coelho, Matthew A. Li, Songyuan Pane, Luna Simona Firth, Mike Ciotta, Giovanni Wrigley, Jonathan D. Cuomo, Maria Emanuela Maresca, Marcello Taylor, Benjamin J. M. BMC Biol Methodology Article BACKGROUND: Base Editing is a precise genome editing method that uses a deaminase-Cas9 fusion protein to mutate cytidine to thymidine in target DNA in situ without the generation of a double-strand break. However, the efficient enrichment of genetically modified cells using this technique is limited by the ability to detect such events. RESULTS: We have developed a Base Editing FLuorescent Activity REporter (BE-FLARE), which allows for the enrichment of cells that have undergone editing of target loci based on a fluorescence shift from BFP to GFP. We used BE-FLARE to evaluate the editing efficiency of APOBEC3A and APOBEC3B family members as alternatives deaminase domains to the rat APOBEC1 domain used in base editor 3 (BE3). We identified human APOBEC3A and APOBEC3B as highly efficient cytidine deaminases for base editing applications with unique properties. CONCLUSIONS: Using BE-FLARE to report on the efficiency and precision of editing events, we outline workflows for the accelerated generation of genetically engineered cell models and the discovery of alternative base editors. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12915-018-0617-1) contains supplementary material, which is available to authorized users. BioMed Central 2018-12-28 /pmc/articles/PMC6309101/ /pubmed/30593278 http://dx.doi.org/10.1186/s12915-018-0617-1 Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Methodology Article Coelho, Matthew A. Li, Songyuan Pane, Luna Simona Firth, Mike Ciotta, Giovanni Wrigley, Jonathan D. Cuomo, Maria Emanuela Maresca, Marcello Taylor, Benjamin J. M. BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title | BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title_full | BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title_fullStr | BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title_full_unstemmed | BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title_short | BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B |
title_sort | be-flare: a fluorescent reporter of base editing activity reveals editing characteristics of apobec3a and apobec3b |
topic | Methodology Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6309101/ https://www.ncbi.nlm.nih.gov/pubmed/30593278 http://dx.doi.org/10.1186/s12915-018-0617-1 |
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