Title |
Genome editing of Clostridium autoethanogenum using CRISPR/Cas9
|
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Published in |
Biotechnology for Biofuels and Bioproducts, October 2016
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DOI | 10.1186/s13068-016-0638-3 |
Pubmed ID | |
Authors |
Shilpa Nagaraju, Naomi Kathleen Davies, David Jeffrey Fraser Walker, Michael Köpke, Séan Dennis Simpson |
Abstract |
Impactful greenhouse gas emissions abatement can now be achieved through gas fermentation using acetogenic microbes for the production of low-carbon fuels and chemicals. However, compared to traditional hosts like Escherichia coli or yeast, only basic genetic tools exist for gas-fermenting acetogens. To advance the process, a robust genetic engineering platform for acetogens is essential. In this study, we report scarless genome editing of an industrially used model acetogen, Clostridium autoethanogenum, using the CRISPR/Cas9 system. Initial efforts to retrofit the CRISPR/Cas9 system for C. autoethanogenum resulted in poor efficiency likely due to uncontrolled expression of Cas9. To address this, we constructed and screened a small library of tetracycline-inducible promoters that can also be used to fine-tune gene expression. With a new inducible promoter, the efficiency of CRISPR/Cas9-mediated desired gene deletion in C. autoethanogenum was improved to over 50 %, making it a viable tool for engineering C. autoethanogenum. Addition of both an inducible promoter library and a scarless genome editing tool is an important expansion to the genetic tool box of industrial C. autoethanogenum strain. |
X Demographics
Geographical breakdown
Country | Count | As % |
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United States | 3 | 27% |
Spain | 2 | 18% |
United Kingdom | 1 | 9% |
Netherlands | 1 | 9% |
Guinea | 1 | 9% |
Unknown | 3 | 27% |
Demographic breakdown
Type | Count | As % |
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Members of the public | 5 | 45% |
Scientists | 4 | 36% |
Science communicators (journalists, bloggers, editors) | 1 | 9% |
Practitioners (doctors, other healthcare professionals) | 1 | 9% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
United Kingdom | 1 | <1% |
New Zealand | 1 | <1% |
Unknown | 213 | 99% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Ph. D. Student | 44 | 20% |
Researcher | 41 | 19% |
Student > Master | 25 | 12% |
Student > Bachelor | 19 | 9% |
Student > Doctoral Student | 10 | 5% |
Other | 28 | 13% |
Unknown | 48 | 22% |
Readers by discipline | Count | As % |
---|---|---|
Biochemistry, Genetics and Molecular Biology | 70 | 33% |
Agricultural and Biological Sciences | 45 | 21% |
Chemical Engineering | 12 | 6% |
Engineering | 10 | 5% |
Immunology and Microbiology | 9 | 4% |
Other | 13 | 6% |
Unknown | 56 | 26% |