ArticleJournal of biological engineering2012
Computational design and characterization of a temperature-sensitive plasmid replicon for gram positive thermophiles.
Article in Journal of biological engineering, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
What it found
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Who cites it
18 citing papers in PubMed, 35 citations in OpenAlex.
- Temperature-dependent dynamics of DNA methylation and plasmid copy number in Yersinia pestis.BMC genomics · 2026Article
- An Initial Genome Editing Toolset for Caldimonas thermodepolymerans, the First Model of Thermophilic Polyhydroxyalkanoates Producer.Microbial biotechnology · 2025Article
- Laboratory Evolution and Reverse Engineering ofApplied and environmental microbiology · 2021Article
- Coculture with hemicellulose-fermenting microbes reverses inhibition of corn fiber solubilization by Clostridium thermocellum at elevated solids loadings.Biotechnology for biofuels · 2021Article
- Development of both type I-B and type II CRISPR/Cas genome editing systems in the cellulolytic bacteriumMetabolic engineering communications · 2020Article
- Conversion of phosphoenolpyruvate to pyruvate inMetabolic engineering communications · 2020Article
- Deletion of the Clostridium thermocellum recA gene reveals that it is required for thermophilic plasmid replication but not plasmid integration at homologous DNA sequences.Journal of industrial microbiology & biotechnology · 2018Article
- Genetic Tools and Techniques for Recombinant Expression in Thermophilic Bacillaceae.Microorganisms · 2018Review
- Expressing theBiotechnology for biofuels · 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
- Development and implementation of rapid metabolic engineering tools for chemical and fuel production inBiotechnology for biofuels · 2017Article
- Progress in understanding and overcoming biomass recalcitrance: a BioEnergy Science Center (BESC) perspective.Biotechnology for biofuels · 2017Article
- Promiscuous plasmid replication in thermophiles: Use of a novel hyperthermophilic replicon for genetic manipulation ofMetabolic engineering communications · 2016Article
- Development of a plasmid-based expression system inMetabolic engineering communications · 2016Article
- Identifying promoters for gene expression inMetabolic engineering communications · 2015Article
- Physiological roles of pyruvate ferredoxin oxidoreductase and pyruvate formate-lyase in Thermoanaerobacterium saccharolyticum JW/SL-YS485.Biotechnology for biofuels · 2015Article
- The emergence of Clostridium thermocellum as a high utility candidate for consolidated bioprocessing applications.Frontiers in chemistry · 2014Review
- Atypical glycolysis in Clostridium thermocellum.Applied and environmental microbiology · 2013Article
Corrections and comments
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Authors and funding
2 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundTemperature-sensitive (Ts) plasmids are useful tools for genetic engineering, but there are currently none compatible with the gram positive, thermophilic, obligate anaerobe, Clostridium thermocellum. Traditional mutagenesis techniques yield Ts mutants at a low frequency, and therefore requires the development of high-throughput screening protocols, which are also not available for this organism. Recently there has been progress in the development of computer algorithms which can predict Ts mutations. Most plasmids currently used for genetic modification of C. thermocellum are based on the replicon of plasmid pNW33N, which replicates using the RepB replication protein. To address this problem, we set out to create a Ts plasmid by mutating the gene coding for the RepB replication protein using an algorithm designed by Varadarajan et al. (1996) for predicting Ts mutants based on the amino-acid sequence of the protein.
resultsA library of 34 mutant plasmids was designed, synthesized and screened, resulting in 6 mutants which exhibited a Ts phenotype. Of these 6, the one with the most temperature-sensitive phenotype (M166A) was compared with the original plasmid. It exhibited lower stability at 48°C and was completely unable to replicate at 55°C.
conclusionsThe plasmid described in this work could be useful in future efforts to genetically engineer C. thermocellum, and the method used to generate this plasmid may be useful for others trying to make Ts plasmids.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.