Project description:Butanol is a bulk chemical feedstock and a promising fuels. Microbial production of butanol is challenging primarily because of its toxicity and low titer of production. Transcript regulator factor Rob plays an important role in butanol-tolerant functional revealed by our previous study. In this study, the mutant strain DTrob (AT686-687 deletion in rob gene) could tolerate 1.25 %(v/v) butanol. And the per unit intracellular butanol concentration and transcriptome of wild-type and DTrob were further compared to understand the regulation mechanism of Rob for butanol tolerance. A total of 285 DEGs (differentially expression genes) were identified by bioinformatic analysis, and they may function in transport and localization according to the GO functional annotations and the functional annotation of gene product in NCBI database. Eighteen DEGs representing different functional categories were selected for real-time quantitative PCR (qPCR) for confirming the dependability of RNA-seq data. The deletion strains of glgS or yibT gene, which were down-regulated by Rob, had butanol-toleranct characteristic, indicating that the deletion or downregulated expression of the two genes down-regulated by Rob could improve the butanol tolerance. This study reveals the transcript regulator factor Rob could regulate and control the expression of the butanol tolerant genes, and its inactivation would result in an improved butanol tolerance.
Project description:The goal of this study is to explore genes that are differentially expressed in E. coli C strains (wt and a butanol-tolerant mutant) after 1-butanol treatment. The butanol-tolerant mutant strain PKH5000 (denoted by 'E' for 'evolved') were derived from KCTC 2571 (wt) (denoted by 'A' for 'ancestral') by proton beam irradiation. 0 and 1 in sample title mean before and after butanol treatment, respectively.
Project description:n-Butanol has been proposed as an alternative biofuel to ethanol, and both Escherichia coli and Saccharomyces cerevisiae have been engineered to produce it. Unfortunately, n-butanol is more toxic than ethanol to these organisms. To understand the basis for its toxicity, cell wide studies were conducted at the transcript, protein and metabolite levels to obtain a global view of the n-butanol stress response. Analysis of the data indicate that n-butanol stress has components common to other stress responses and includes perturbation in respiratory functions (nuo, cyo operons), oxidative stress (sodC, katG, yqhD), heat shock and cell envelope stress (rpoE, clpB, htpG, degP, cpxPR), metabolite transport (malE, opp operon) and biosynthesis. Inducible expression of the yqhD gene was found to improve the host’s tolerance to exogenous n-butanol and confirms the role of this gene in coping with butanol stress. To survey for other potential candidates that may serve to improve host tolerance, mutant strains in several candidates which show changes at the transcript and protein levels were examined for sensitivity during butanol exposure. Chassis engineering based on these cues may be required in a high production titer, butanol-producing host.
Project description:Metabolite accumulation has pleiotropic, including toxic, effects on cellular physiology, with a variety of genetic resistance mechanisms previously identified. Using random genomic libraries and DNA microarrays, library inserts were preferentially enriched by culturing in media containing butanol, with successive transfers into fresh media containing incrementally increasing butanol concentrations. Keywords: genomic library preferential enrichment
Project description:We report the application of a high-throughput technique, RNA-seq, to study the transcriptomic response of P. putida BIRD1 in the presence of butanol with the aim to study in more detail the mechanisms involved in butanol response. Our work is the first global transcriptomic analysis done in P. putida BIRD1 in the presence of butanol.
Project description:Clostridium acetobutylicum is a Gram positive, endospore forming firmicute that has been known as the model organims for ABE (acetone-butanol-ethanol) fermentation. With its ability to consume a wide variety of substrates, C. acetobutylicum carries out a biphasic ABE fermentation, which consists of the acidogenic growth phase with the formation of butyric acid and acetic acid, followed by the solventogenic stationary phase with the formation of acetone, butanol and ethanol, characterised by the reassimilation of acids. The production butanol is of renewed ineterest both as a potential biofuel and bulk chemical production. Both butanol and butyrate posses toxic characteristic and here, we focus on understanding and modeling the stress response of C. acetobutylicum to one of the two important toxic metabolites: butanol.
Project description:We successfully isolated an E. coli strain harboring rpoD mutant B8 with 2% (v/v) butanol tolerance using global transcriptional machinery engineering approach. DNA microarrays were employed to assess the transcriptome profile of n-butanol tolerance strain B8 and control strain E. coli JM109. The goal of this study is therefore to identify E. coli genes that are involved in n-butanol tolerance.
Project description:Clostridium acetobutylicum is well-known for its butanol production. Butanol toxicity is a major drawback for the generation of high-butanol producing strains. Here, the transcriptional response a steady state, acidogenic (pH 6), phosphate-limited Clostridium acetobutylicum chemostat culture to different levels of n-butanol (0.25-1%) was investigated. For the butanol challenge experiments butanol (1-butanol) was added (a) to the supplying medium and (b) to the culture vessel to guarantee an immediate change in the butanol concentration. Addition of butanol to the culture was timed to match the supply of the new medium through the feedlines. The butanol concentration was increased stepwise in intervals of 66.6 h (5 volume changes) to moderate butanol concentrations of 0.25%, 0.5%, 0.75% and 1% (v/v).
Project description:Genomic Library Enrichment to determine the n-Butanol Tolerant related genes in E. coli. The samples involves a series of batch transfers to increasing concentrations of n-butanol and controls (always grew in absence of the solvent)
Project description:Clostridium acetobutylicum is a Gram positive, endospore forming firmicute that has been known as the model organims for ABE (acetone-butanol-ethanol) fermentation. With its ability to consume a wide variety of substrates, C. acetobutylicum carries out a biphasic ABE fermentation, which consists of the acidogenic growth phase with the formation of butyric acid and acetic acid, followed by the solventogenic stationary phase with the formation of acetone, butanol and ethanol, characterised by the reassimilation of acids. The production butanol is of renewed ineterest both as a potential biofuel and bulk chemical production. Both butanol and butyric acid posses toxic characteristic and here, we focus on understanding and modeling the stress response of C. acetobutylicum to one of the two important toxic metabolites: butyric acid.