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A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection
The use of CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein) for sequence-specific elimination of bacteria or resistance genes is a powerful tool for combating antibiotic resistance. However, this approach requires efficient delivery of CRISPR/Cas DNA c...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
PeerJ Inc.
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8428258/ https://www.ncbi.nlm.nih.gov/pubmed/34567840 http://dx.doi.org/10.7717/peerj.11996 |
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author | Wongpayak, Panjaporn Meesungnoen, Orapan Saejang, Somchai Subsoontorn, Pakpoom |
author_facet | Wongpayak, Panjaporn Meesungnoen, Orapan Saejang, Somchai Subsoontorn, Pakpoom |
author_sort | Wongpayak, Panjaporn |
collection | PubMed |
description | The use of CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein) for sequence-specific elimination of bacteria or resistance genes is a powerful tool for combating antibiotic resistance. However, this approach requires efficient delivery of CRISPR/Cas DNA cassette(s) into the targeted bacterial population. Compared to phage transduction, plasmid conjugation can deliver DNA to a broader host range but often suffers from low delivery efficiency. Here, we developed multi-plasmid conjugation systems for efficient CRISPR/Cas delivery, target DNA elimination and plasmid replacement. The CRISPR/Cas system, delivered via a broad-host-range R1162 mobilizable plasmid, specifically eliminated the targeted plasmid in recipient cells. A self-transmissible RK2 helper plasmid facilitated the spread of mobilizable CRISPR/Cas. The replacement of the target plasmid with another plasmid from the same compatibility group helped speed up target plasmid elimination especially when the target plasmid was also mobilizable. Together, we showed that up to 100% of target plasmid from the entire recipient population could be replaced even at a low (1:180) donor-to-recipient ratio and in the absence of transconjugant selection. Such an ability to modify genetic content of microbiota efficiently in the absence of selection will be critical for future development of CRISPR antimicrobials as well as genetic tools for in situ microbiome engineering. |
format | Online Article Text |
id | pubmed-8428258 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | PeerJ Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-84282582021-09-24 A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection Wongpayak, Panjaporn Meesungnoen, Orapan Saejang, Somchai Subsoontorn, Pakpoom PeerJ Bioengineering The use of CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein) for sequence-specific elimination of bacteria or resistance genes is a powerful tool for combating antibiotic resistance. However, this approach requires efficient delivery of CRISPR/Cas DNA cassette(s) into the targeted bacterial population. Compared to phage transduction, plasmid conjugation can deliver DNA to a broader host range but often suffers from low delivery efficiency. Here, we developed multi-plasmid conjugation systems for efficient CRISPR/Cas delivery, target DNA elimination and plasmid replacement. The CRISPR/Cas system, delivered via a broad-host-range R1162 mobilizable plasmid, specifically eliminated the targeted plasmid in recipient cells. A self-transmissible RK2 helper plasmid facilitated the spread of mobilizable CRISPR/Cas. The replacement of the target plasmid with another plasmid from the same compatibility group helped speed up target plasmid elimination especially when the target plasmid was also mobilizable. Together, we showed that up to 100% of target plasmid from the entire recipient population could be replaced even at a low (1:180) donor-to-recipient ratio and in the absence of transconjugant selection. Such an ability to modify genetic content of microbiota efficiently in the absence of selection will be critical for future development of CRISPR antimicrobials as well as genetic tools for in situ microbiome engineering. PeerJ Inc. 2021-09-06 /pmc/articles/PMC8428258/ /pubmed/34567840 http://dx.doi.org/10.7717/peerj.11996 Text en © 2021 Wongpayak et al. https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited. |
spellingShingle | Bioengineering Wongpayak, Panjaporn Meesungnoen, Orapan Saejang, Somchai Subsoontorn, Pakpoom A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title | A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title_full | A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title_fullStr | A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title_full_unstemmed | A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title_short | A highly effective and self-transmissible CRISPR antimicrobial for elimination of target plasmids without antibiotic selection |
title_sort | highly effective and self-transmissible crispr antimicrobial for elimination of target plasmids without antibiotic selection |
topic | Bioengineering |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8428258/ https://www.ncbi.nlm.nih.gov/pubmed/34567840 http://dx.doi.org/10.7717/peerj.11996 |
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