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Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion

Precise genetic manipulation is vital to studying bacterial physiology, but is difficult to achieve in some bacterial species due to the weak intrinsic homologous recombination (HR) capacity and lack of a compatible exogenous HR system. Here we report the establishment of a rapid and efficient metho...

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Autores principales: Zhang, Ya, Zhang, Hongyuan, Wang, Zhipeng, Wu, Zhaowei, Wang, Yu, Tang, Na, Xu, Xuexia, Zhao, Suwen, Chen, Weizhong, Ji, Quanjiang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Royal Society of Chemistry 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7069399/
https://www.ncbi.nlm.nih.gov/pubmed/32206285
http://dx.doi.org/10.1039/c9sc03784e
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author Zhang, Ya
Zhang, Hongyuan
Wang, Zhipeng
Wu, Zhaowei
Wang, Yu
Tang, Na
Xu, Xuexia
Zhao, Suwen
Chen, Weizhong
Ji, Quanjiang
author_facet Zhang, Ya
Zhang, Hongyuan
Wang, Zhipeng
Wu, Zhaowei
Wang, Yu
Tang, Na
Xu, Xuexia
Zhao, Suwen
Chen, Weizhong
Ji, Quanjiang
author_sort Zhang, Ya
collection PubMed
description Precise genetic manipulation is vital to studying bacterial physiology, but is difficult to achieve in some bacterial species due to the weak intrinsic homologous recombination (HR) capacity and lack of a compatible exogenous HR system. Here we report the establishment of a rapid and efficient method for directly converting adenine to guanine in bacterial genomes using the fusion of an adenine deaminase and a Cas9 nickase. The method achieves the conversion of adenine to guanine via an enzymatic deamination reaction and a subsequent DNA replication process rather than HR, which is utilized in conventional bacterial genetic manipulation methods, thereby substantially simplifying the genome editing process. A systematic screening targeting the possibly editable adenine sites of cntBC, the importer of the staphylopine/metal complex in Staphylococcus aureus, pinpoints key residues for metal importation, demonstrating that application of the system would greatly facilitate the genomic engineering of bacteria.
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spelling pubmed-70693992020-03-23 Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion Zhang, Ya Zhang, Hongyuan Wang, Zhipeng Wu, Zhaowei Wang, Yu Tang, Na Xu, Xuexia Zhao, Suwen Chen, Weizhong Ji, Quanjiang Chem Sci Chemistry Precise genetic manipulation is vital to studying bacterial physiology, but is difficult to achieve in some bacterial species due to the weak intrinsic homologous recombination (HR) capacity and lack of a compatible exogenous HR system. Here we report the establishment of a rapid and efficient method for directly converting adenine to guanine in bacterial genomes using the fusion of an adenine deaminase and a Cas9 nickase. The method achieves the conversion of adenine to guanine via an enzymatic deamination reaction and a subsequent DNA replication process rather than HR, which is utilized in conventional bacterial genetic manipulation methods, thereby substantially simplifying the genome editing process. A systematic screening targeting the possibly editable adenine sites of cntBC, the importer of the staphylopine/metal complex in Staphylococcus aureus, pinpoints key residues for metal importation, demonstrating that application of the system would greatly facilitate the genomic engineering of bacteria. Royal Society of Chemistry 2020-01-06 /pmc/articles/PMC7069399/ /pubmed/32206285 http://dx.doi.org/10.1039/c9sc03784e Text en This journal is © The Royal Society of Chemistry 2020 http://creativecommons.org/licenses/by-nc/3.0/ This article is freely available. This article is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported Licence (CC BY-NC 3.0)
spellingShingle Chemistry
Zhang, Ya
Zhang, Hongyuan
Wang, Zhipeng
Wu, Zhaowei
Wang, Yu
Tang, Na
Xu, Xuexia
Zhao, Suwen
Chen, Weizhong
Ji, Quanjiang
Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title_full Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title_fullStr Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title_full_unstemmed Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title_short Programmable adenine deamination in bacteria using a Cas9–adenine-deaminase fusion
title_sort programmable adenine deamination in bacteria using a cas9–adenine-deaminase fusion
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7069399/
https://www.ncbi.nlm.nih.gov/pubmed/32206285
http://dx.doi.org/10.1039/c9sc03784e
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