ArticleBiotechnology for biofuels2016
A markerless gene deletion and integration system for Thermoanaerobacter ethanolicus.
Article in Biotechnology for biofuels, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 32 citations in OpenAlex.
- Adaptive evolution of electron transfer pathways inJournal of bacteriology · 2026Article
- A thermostable Cas9-based genome editing system for thermophilic acetogenic bacteriumApplied and environmental microbiology · 2025Article
- Identification of electron transfer enzymes inJournal of bacteriology · 2025Article
- Metabolic engineering of Thermoanaerobacterium AK17 for increased ethanol production in seaweed hydrolysate.Biotechnology for biofuels and bioproducts · 2023Article
- Exploiting the Type I-B CRISPR Genome Editing System in Thermoanaerobacterium aotearoense SCUT27 and Engineering the Strain for Enhanced Ethanol Production.Applied and environmental microbiology · 2022Article
- Alcohol dehydrogenases AdhE and AdhB with broad substrate ranges are important enzymes for organic acid reduction in Thermoanaerobacter sp. strain X514.Biotechnology for biofuels · 2021Article
- Genome Editing of the Anaerobic Thermophile Thermoanaerobacter ethanolicus Using Thermostable Cas9.Applied and environmental microbiology · 2020Article
- A Hybrid Sequencing Approach Completes the Genome Sequence of Thermoanaerobacter ethanolicus JW 200.Microbiology resource announcements · 2019Article
- A Genetic System for the Thermophilic Acetogenic Bacterium Thermoanaerobacter kivui.Applied and environmental microbiology · 2018Article
- Determining the roles of the three alcohol dehydrogenases (AdhA, AdhB and AdhE) in Thermoanaerobacter ethanolicus during ethanol formation.Journal of industrial microbiology & biotechnology · 2017Article
- BothJournal of bacteriology · 2017Article
- "Hot" acetogenesis.Extremophiles : life under extreme conditions · 2017Review
- Ferredoxin:NAD+ Oxidoreductase of Thermoanaerobacterium saccharolyticum and Its Role in Ethanol Formation.Applied and environmental microbiology · 2016Article
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Authors and funding
4 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundThermoanaerobacter ethanolicus produces a considerable amount of ethanol from a range of carbohydrates and is an attractive candidate for applications in bioconversion processes. A genetic system with reusable selective markers would be useful for deleting acid production pathways as well as other genetic modifications.
resultsThe thymidine kinase (tdk) gene was deleted from T. ethanolicus JW200 to allow it to be used as a selectable marker, resulting in strain X20. Deletion of the tdk gene reduced growth rate by 20 %; however, this could be reversed by reintroducing the tdk gene (strain X20C). The tdk and high-temperature kanamycin (htk) markers were tested by using them to delete lactate dehydrogenase (ldh). During positive selection of ldh knockouts in strain X20 on kanamycin agar plates, six out of seven picked colonies were verified transformants. Deletion of ldh reduced lactic acid production by 90 %. The tdk and 5-fluoro-2'-deoxyuridine (FUDR) combination worked reliably as demonstrated by successful tdk removal in all 21 colonies tested.
conclusionA gene deletion and integration system with reusable markers has been developed for Thermoanaerobacter ethanolicus JW200 with positive selection on kanamycin and negative selection on FUDR. Gene deletion was demonstrated by ldh gene deletion and gene integration was demonstrated by re-integration of the tdk gene. Transformation via a natural competence protocol could use DNA PCR products amplified directly from Gibson Assembly mixture for efficient genetic modification.
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