ArticleBiotechnology for biofuels2016
Dam and Dcm methylations prevent gene transfer into Clostridium pasteurianum NRRL B-598: development of methods for electrotransformation, conjugation, and sonoporation.
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 17 papers.
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17 citing papers in PubMed.
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- Multiplexed Assembly and Annotation of Synthetic Biology Constructs Using Long-Read Nanopore Sequencing.ACS synthetic biology · 2022Article
- Novel thermophilic polyhydroxyalkanoates producing strain Aneurinibacillus thermoaerophilus CCM 8960.Applied microbiology and biotechnology · 2022Article
- Breaking the Restriction Barriers and Applying CRISPRi as a Gene Silencing Tool inMicroorganisms · 2022Article
- Fluorescent Protein Expression as a Proxy for Bacterial Fitness in a High-Throughput Assay.Applied and environmental microbiology · 2021Article
- Genetic and metabolic engineering challenges of C1-gas fermenting acetogenic chassis organisms.FEMS microbiology reviews · 2021Review
- Approaches to genetic tool development for rapid domestication of non-model microorganisms.Biotechnology for biofuels · 2021Review
- Changes in efflux pump activity of Clostridium beijerinckii throughout ABE fermentation.Applied microbiology and biotechnology · 2021Article
- Article
- σScientific reports · 2019Article
- Acidogenesis, solventogenesis, metabolic stress response and life cycle changes in Clostridium beijerinckii NRRL B-598 at the transcriptomic level.Scientific reports · 2019Article
- Article
- Development of a metabolic pathway transfer and genomic integration system for the syngas-fermenting bacteriumBiotechnology for biofuels · 2019Article
- A transcriptional response ofBiotechnology for biofuels · 2019Article
- A novel method for transforming the thermophilic bacterium Geobacillus kaustophilus.Microbial cell factories · 2018Article
- A roadmap for gene system development in Clostridium.Anaerobe · 2016Review
- Disruption of the Reductive 1,3-Propanediol Pathway Triggers Production of 1,2-Propanediol for Sustained Glycerol Fermentation by Clostridium pasteurianum.Applied and environmental microbiology · 2016Article
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4 authors.
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Abstract
backgroundButanol is currently one of the most discussed biofuels. Its use provides many benefits in comparison to bio-ethanol, but the price of its fermentative production is still high. Genetic improvements could help solve many problems associated with butanol production during ABE fermentation, such as its toxicity, low concentration achievable in the cultivation medium, the need for a relatively expensive substrate, and many more. Clostridium pasteurianum NRRL B-598 is non-type strain producing butanol, acetone, and a negligible amount of ethanol. Its main benefits are high oxygen tolerance, utilization of a wide range of carbon and nitrogen sources, and the availability of its whole genome sequence. However, there is no established method for the transfer of foreign DNA into this strain; this is the next step necessary for progress in its use for butanol production.
resultsWe have described functional protocols for conjugation and transformation of the bio-butanol producer C. pasteurianum NRRL B-598 by foreign plasmid DNA. We show that the use of unmethylated plasmid DNA is necessary for efficient transformation or successful conjugation. Genes encoding DNA methylation and those for restriction-modification systems and antibiotic resistance were searched for in the whole genome sequence and their homologies with other clostridial bacteria were determined. Furthermore, activity of described novel type I restriction system was proved experimentally. The described electrotransformation protocol achieved an efficiency 1.2 × 10(2) cfu/μg DNA after step-by-step optimization and an efficiency of 1.6 × 10(2) cfu/μg DNA was achieved by the sonoporation technique using a standard laboratory ultrasound bath. The highest transformation efficiency was achieved using a combination of these approaches; sono/electroporation led to an increase in transformation efficiency, to 5.3 × 10(2) cfu/μg DNA.
conclusionsBoth Dam and Dcm methylations are detrimental for transformation of C. pasteurianum NRRL B-598. Methods for conjugation, electroporation, sonoporation, and a combined method for sono/electroporation were established for this strain. The methods described could be used for genetic improvement of this strain, which is suitable for bio-butanol production.
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