Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli
Summary: Plasmids are the workhorse of both industrial biotechnology and synthetic biology, but ensuring they remain in bacterial cells is a challenge. Antibiotic selection cannot be used to stabilize plasmids in most real-world applications, and inserting dynamical gene networks into the genome rem...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2019-04-01
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| Series: | iScience |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2589004219300859 |
| _version_ | 1818525346611331072 |
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| author | Alex J.H. Fedorec Tanel Ozdemir Anjali Doshi Yan-Kay Ho Luca Rosa Jack Rutter Oscar Velazquez Vitor B. Pinheiro Tal Danino Chris P. Barnes |
| author_facet | Alex J.H. Fedorec Tanel Ozdemir Anjali Doshi Yan-Kay Ho Luca Rosa Jack Rutter Oscar Velazquez Vitor B. Pinheiro Tal Danino Chris P. Barnes |
| author_sort | Alex J.H. Fedorec |
| collection | DOAJ |
| description | Summary: Plasmids are the workhorse of both industrial biotechnology and synthetic biology, but ensuring they remain in bacterial cells is a challenge. Antibiotic selection cannot be used to stabilize plasmids in most real-world applications, and inserting dynamical gene networks into the genome remains challenging. Plasmids have evolved several mechanisms for stability, one of which, post-segregational killing (PSK), ensures that plasmid-free cells do not survive. Here we demonstrate the plasmid-stabilizing capabilities of the axe/txe toxin-antitoxin system and the microcin-V bacteriocin system in the probiotic bacteria Escherichia coli Nissle 1917 and show that they can outperform the commonly used hok/sok. Using plasmid stability assays, automated flow cytometry analysis, mathematical models, and Bayesian statistics we quantified plasmid stability in vitro. Furthermore, we used an in vivo mouse cancer model to demonstrate plasmid stability in a real-world therapeutic setting. These new PSK systems, plus the developed Bayesian methodology, will have wide applicability in clinical and industrial biotechnology. : Biological Sciences; Gene Network; Bioengineering Subject Areas: Biological Sciences, Gene Network, Bioengineering |
| first_indexed | 2024-12-11T06:08:11Z |
| format | Article |
| id | doaj.art-7fc97d305d384cc1b919e95418a68bcc |
| institution | Directory Open Access Journal |
| issn | 2589-0042 |
| language | English |
| last_indexed | 2024-12-11T06:08:11Z |
| publishDate | 2019-04-01 |
| publisher | Elsevier |
| record_format | Article |
| series | iScience |
| spelling | doaj.art-7fc97d305d384cc1b919e95418a68bcc2022-12-22T01:18:13ZengElsevieriScience2589-00422019-04-0114323334Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coliAlex J.H. Fedorec0Tanel Ozdemir1Anjali Doshi2Yan-Kay Ho3Luca Rosa4Jack Rutter5Oscar Velazquez6Vitor B. Pinheiro7Tal Danino8Chris P. Barnes9Department of Cell and Developmental Biology, University College London, London WC1E 6BT, UK; Centre for Mathematics, Physics and Engineering in the Life Sciences and Experimental Biology, University College London, London WC1E 6BT, UK; Corresponding authorDepartment of Cell and Developmental Biology, University College London, London WC1E 6BT, UKDepartment of Biomedical Engineering, Columbia University, New York City, NY 10027, USAInstitute of Structural and Molecular Biology, University College London, London WC1E 6BT, UKDepartment of Cell and Developmental Biology, University College London, London WC1E 6BT, UKDepartment of Cell and Developmental Biology, University College London, London WC1E 6BT, UKDepartment of Biomedical Engineering, Columbia University, New York City, NY 10027, USAInstitute of Structural and Molecular Biology, University College London, London WC1E 6BT, UK; KU Leuven Rega Institute for Medical Research, Herestraat, 49 Box 1030, 3000 Leuven, BelgiumDepartment of Biomedical Engineering, Columbia University, New York City, NY 10027, USA; Data Science Institute, Columbia University, New York, NY 10027, USA; Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY 10032, USADepartment of Cell and Developmental Biology, University College London, London WC1E 6BT, UK; Department of Genetics, Evolution and Environment, University College London, London WC1E 6BT, UK; Corresponding authorSummary: Plasmids are the workhorse of both industrial biotechnology and synthetic biology, but ensuring they remain in bacterial cells is a challenge. Antibiotic selection cannot be used to stabilize plasmids in most real-world applications, and inserting dynamical gene networks into the genome remains challenging. Plasmids have evolved several mechanisms for stability, one of which, post-segregational killing (PSK), ensures that plasmid-free cells do not survive. Here we demonstrate the plasmid-stabilizing capabilities of the axe/txe toxin-antitoxin system and the microcin-V bacteriocin system in the probiotic bacteria Escherichia coli Nissle 1917 and show that they can outperform the commonly used hok/sok. Using plasmid stability assays, automated flow cytometry analysis, mathematical models, and Bayesian statistics we quantified plasmid stability in vitro. Furthermore, we used an in vivo mouse cancer model to demonstrate plasmid stability in a real-world therapeutic setting. These new PSK systems, plus the developed Bayesian methodology, will have wide applicability in clinical and industrial biotechnology. : Biological Sciences; Gene Network; Bioengineering Subject Areas: Biological Sciences, Gene Network, Bioengineeringhttp://www.sciencedirect.com/science/article/pii/S2589004219300859 |
| spellingShingle | Alex J.H. Fedorec Tanel Ozdemir Anjali Doshi Yan-Kay Ho Luca Rosa Jack Rutter Oscar Velazquez Vitor B. Pinheiro Tal Danino Chris P. Barnes Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli iScience |
| title | Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli |
| title_full | Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli |
| title_fullStr | Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli |
| title_full_unstemmed | Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli |
| title_short | Two New Plasmid Post-segregational Killing Mechanisms for the Implementation of Synthetic Gene Networks in Escherichia coli |
| title_sort | two new plasmid post segregational killing mechanisms for the implementation of synthetic gene networks in escherichia coli |
| url | http://www.sciencedirect.com/science/article/pii/S2589004219300859 |
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