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Transcriptome analysis of isolated Escherichia coli mutants using VERT for the study of n-butanol tolerance

ABSTRACT: Transcriptome analysis of isolated mutants using the method Visualizing Evolution in Real-Time (VERT) for the study of n-butanol tolerance. The samples were isolated from an evolution experiment picking samples at different times based in the evolution dynamics obtained with VERT. Mutants were grown in chemostats at 0.8% (v/v) of n-butanol and compared with the expression of wild-type to the same concentration of solvent.

ORGANISM(S): Escherichia coli K-12 Escherichia coli

PROVIDER: GSE30005 | GEO | 2012/03/31

SECONDARY ACCESSION(S): PRJNA143677

REPOSITORIES: GEO

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Transcriptome analysis of isolated Escherichia coli mutants using VERT for the study of n-butanol tolerance

Project description:Transcriptome analysis of isolated mutants using the method Visualizing Evolution in Real-Time (VERT) for the study of n-butanol tolerance. The samples were isolated from an evolution experiment picking samples at different times based in the evolution dynamics obtained with VERT. Mutants were grown in chemostats at 0.8% (v/v) of n-butanol and compared with the expression of wild-type to the same concentration of solvent. Three biological replicas. Each sample represents an isolated mutant. The reference for each mutant corresponds to wild-type, both exposed to the same concentration of n-butanol (0.8% (v/v)).

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CcpA, a Pleiotropic Key Regulator in Butanol-producing Clostridium acetobutylicum ATCC 824

Project description:Clostridium acetobutylicum is a typical bacterium of major importance to industrial butanol production. In order to dissect the regulatory network pertaining to the industrial application of this bacterium, catabolite control protein A (CcpA) was investigated for its global function by DNA microarray.It showed that CcpA of C. acetobutylicum controls hundreds of genes, not only carbon metabolism, but also solvent production and sporulation in the life cycle.The results here demonstrated that CcpA is an important pleiotropic regulator related to some specific physiological and biochemical process in butanol-producing C. acetobutylicum.

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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 hostâs 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. This comparison is between E. coli DH1 cells treated with 0.8% n-butanol and untreated cells at 0, 30, 80, and 195 minutes after addition. 3 biological replicates were grown, total RNA was extracted, labeled, and hybridized on 3 slides for each time point. http://www.microbesonline.org/cgi-bin/microarray/viewExp.cgi?expId=1266

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Transcriptional Response of the thermoacidophilic Archaeon Sulfolobus acidocaldarius to solvent stress exemplified by 1-butanol exposure

Project description:To study the cellular global response of the thermoacidophilic Archaeon Sulfolobus acidocaldarius upon solvent exposure we performed transcriptome comparing the response of planktonic and biofilm cells in absence and presence 1-butanol. S. acidocaldarius was grown in Petri dishes (static incubation) in the absence and presence of 0.5% and 1% (v/v) 1-butanol at 76°C for four days. Planktonic and biofilm cells were harvested and the transcriptional response towards 1-butanol was analysed via RNA-seq. In detail:Biofilm and planktonic cell samples (supernatant after static incubation) were generated in Petri dishes (92 x 16 mm, Polystyrene, without cams, Sarstedt) as described before. Cells grown under six different conditions, including Biofilm-Control (BF0), Biofilm-0.5% (v/v) 1-Butanol (BF05), Biofilm-1% (v/v) 1-Butanol (BF1), Planktonic-Control (PL0), Planktonic-0.5% (v/v) 1-Butanol (PL05), Planktonic-1% (v/v) 1-Butanol (PL1), were seperated, harvested by centrifugation (10 min, 5,000 x g, 4 °C) and immediately frozen at -70 °C. Isolation of cells was performed in triplicates with ten technical replicas each. Samples were pooled to obtain sufficient cell mass for further processing. RNA was isolated using Trizol (Thermo Fisher Scientific). In accordance, sequencing libraries were prepared via Illumina TruSeq stranded mRNA using Ribozero for RNA depletion. Sequencing was performed on a MiSeq instrument (Illumina, San Diego, California, USA) using v3 chemistry with a read length of 2x76 nt.

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