A roadmap for gene system development in Clostridium
Clostridium species are both heroes and villains. Some cause serious human and animal diseases, those present in the microbiota contribute to health and wellbeing, while others represent useful industrial chassis for the production of chemicals and fuels. To understand, counter or exploit, there is...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
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Elsevier
2016
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| Online Access: | https://eprints.nottingham.ac.uk/35934/ |
| _version_ | 1848795193244909568 |
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| author | Minton, Nigel P. Ehsaan, Muhammad Humphreys, Christopher M. Little, Gareth T. Baker, Jonathan Henstra, Anne M. Liew, Fungmin Kelly, Michelle Sheng, Lili Schwarz, Katrin Zhang, Ying |
| author_facet | Minton, Nigel P. Ehsaan, Muhammad Humphreys, Christopher M. Little, Gareth T. Baker, Jonathan Henstra, Anne M. Liew, Fungmin Kelly, Michelle Sheng, Lili Schwarz, Katrin Zhang, Ying |
| author_sort | Minton, Nigel P. |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | Clostridium species are both heroes and villains. Some cause serious human and animal diseases, those present in the microbiota contribute to health and wellbeing, while others represent useful industrial chassis for the production of chemicals and fuels. To understand, counter or exploit, there is a fundamental requirement for effective systems that may be used for directed or random genome modifications. We have formulated a simple roadmap whereby the necessary gene systems maybe developed and deployed. At its heart is the use of 'pseudo-suicide' vectors and the creation of a pyrE mutant (a uracil auxotroph), initially aided by ClosTron technology, but ultimately made using a special form of allelic exchange termed ACE (Allele-Coupled Exchange). All mutants, regardless of the mutagen employed, are made in this host. This is because through the use of ACE vectors, mutants can be rapidly complemented concomitant with correction of the pyrE allele and restoration of uracil prototrophy. This avoids the phenotypic effects frequently observed with high copy number plasmids and dispenses with the need to add antibiotic to ensure plasmid retention. Once available, the pyrE host may be used to stably insert all manner of application specific modules. Examples include, a sigma factor to allow deployment of a mariner transposon, hydrolases involved in biomass deconstruction and therapeutic genes in cancer delivery vehicles. To date, provided DNA transfer is obtained, we have not encountered any clostridial species where this technology cannot be applied. These include, Clostridium difficile, Clostridium acetobutylicum, Clostridium beijerinckii, Clostridium botulinum, Clostridium perfringens, Clostridium sporogenes, Clostridium pasteurianum, Clostridium ljungdahlii, Clostridium autoethanogenum and even Geobacillus thermoglucosidasius. |
| first_indexed | 2025-11-14T19:28:12Z |
| format | Article |
| id | nottingham-35934 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| last_indexed | 2025-11-14T19:28:12Z |
| publishDate | 2016 |
| publisher | Elsevier |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-359342020-05-04T18:08:30Z https://eprints.nottingham.ac.uk/35934/ A roadmap for gene system development in Clostridium Minton, Nigel P. Ehsaan, Muhammad Humphreys, Christopher M. Little, Gareth T. Baker, Jonathan Henstra, Anne M. Liew, Fungmin Kelly, Michelle Sheng, Lili Schwarz, Katrin Zhang, Ying Clostridium species are both heroes and villains. Some cause serious human and animal diseases, those present in the microbiota contribute to health and wellbeing, while others represent useful industrial chassis for the production of chemicals and fuels. To understand, counter or exploit, there is a fundamental requirement for effective systems that may be used for directed or random genome modifications. We have formulated a simple roadmap whereby the necessary gene systems maybe developed and deployed. At its heart is the use of 'pseudo-suicide' vectors and the creation of a pyrE mutant (a uracil auxotroph), initially aided by ClosTron technology, but ultimately made using a special form of allelic exchange termed ACE (Allele-Coupled Exchange). All mutants, regardless of the mutagen employed, are made in this host. This is because through the use of ACE vectors, mutants can be rapidly complemented concomitant with correction of the pyrE allele and restoration of uracil prototrophy. This avoids the phenotypic effects frequently observed with high copy number plasmids and dispenses with the need to add antibiotic to ensure plasmid retention. Once available, the pyrE host may be used to stably insert all manner of application specific modules. Examples include, a sigma factor to allow deployment of a mariner transposon, hydrolases involved in biomass deconstruction and therapeutic genes in cancer delivery vehicles. To date, provided DNA transfer is obtained, we have not encountered any clostridial species where this technology cannot be applied. These include, Clostridium difficile, Clostridium acetobutylicum, Clostridium beijerinckii, Clostridium botulinum, Clostridium perfringens, Clostridium sporogenes, Clostridium pasteurianum, Clostridium ljungdahlii, Clostridium autoethanogenum and even Geobacillus thermoglucosidasius. Elsevier 2016-10-01 Article PeerReviewed Minton, Nigel P., Ehsaan, Muhammad, Humphreys, Christopher M., Little, Gareth T., Baker, Jonathan, Henstra, Anne M., Liew, Fungmin, Kelly, Michelle, Sheng, Lili, Schwarz, Katrin and Zhang, Ying (2016) A roadmap for gene system development in Clostridium. Anaerobe, 41 . pp. 104-112. ISSN 1095-8274 Restriction modification; gene transfer; ClosTron; allelic exchange; counterselection marker; pyrE; knock-out; knock-in; Fluoroorotic acid http://www.sciencedirect.com/science/article/pii/S1075996416300646 doi:10.1016/j.anaerobe.2016.05.011 doi:10.1016/j.anaerobe.2016.05.011 |
| spellingShingle | Restriction modification; gene transfer; ClosTron; allelic exchange; counterselection marker; pyrE; knock-out; knock-in; Fluoroorotic acid Minton, Nigel P. Ehsaan, Muhammad Humphreys, Christopher M. Little, Gareth T. Baker, Jonathan Henstra, Anne M. Liew, Fungmin Kelly, Michelle Sheng, Lili Schwarz, Katrin Zhang, Ying A roadmap for gene system development in Clostridium |
| title | A roadmap for gene system development in Clostridium |
| title_full | A roadmap for gene system development in Clostridium |
| title_fullStr | A roadmap for gene system development in Clostridium |
| title_full_unstemmed | A roadmap for gene system development in Clostridium |
| title_short | A roadmap for gene system development in Clostridium |
| title_sort | roadmap for gene system development in clostridium |
| topic | Restriction modification; gene transfer; ClosTron; allelic exchange; counterselection marker; pyrE; knock-out; knock-in; Fluoroorotic acid |
| url | https://eprints.nottingham.ac.uk/35934/ https://eprints.nottingham.ac.uk/35934/ https://eprints.nottingham.ac.uk/35934/ |