ABSTRACT: Expression data from batch cultivations with the acidogenic Aspergillus niger wild type strain ATCC 1015 and the areB complete deletion strain constructed using ATCC 1015 as background strain with glucose or glycerol as carbon sources.
Project description:Expression data from batch cultivations of Aspergillus niger wild type strain ATCC 1015 and adrA, facB and creA deletion mutants constructed on ATCC 1015 background strain with glucose or glycerol as carbon sources. Genome-wide transcriptome analysis was used to identify genes either affected directly or indirectly by each transcription factor investigated during growth on a repressing or a derepressing carbon source. For this purpose, batch cultivations under well-controlled conditions were performed with Aspergillus niger wild type strain ATCC 1015 and the three deletion mutants of the corresponding transcription factors AdrA, FacB and CreA. Samples for RNA extraction were collected and further processed for hybridization in custom-designed Affymetrix microarrays containing probes for three Aspergillus species, including A. niger.
Project description:Expression data from batch cultivations of Aspergillus niger wild type strain ATCC 1015 and adrA, facB and creA deletion mutants constructed on ATCC 1015 background strain with glucose or glycerol as carbon sources. Genome-wide transcriptome analysis was used to identify genes either affected directly or indirectly by each transcription factor investigated during growth on a repressing or a derepressing carbon source. For this purpose, batch cultivations under well-controlled conditions were performed with Aspergillus niger wild type strain ATCC 1015 and the three deletion mutants of the corresponding transcription factors AdrA, FacB and CreA. Samples for RNA extraction were collected and further processed for hybridization in custom-designed Affymetrix microarrays containing probes for three Aspergillus species, including A. niger. Triplicate batch fermentations of each of the four Aspergillus niger strains used, the wild type A. niger strain ATCC 1015 and three gene deletion mutants, were carried out using glucose or glycerol as carbon source, and transcriptome analysis was performed. Biomass from each batch cultivation was harvested in the exponential phase of growth and further processed for RNA extraction and hybridization on Affymetrix microarrays.
Project description:This approach aims at searching unidentified regulatory roles of the AreB transcription factor in the overall carbon metabolism of A. niger. A full areB gene deletion mutant was constructed and characterized in A. niger ATCC 1015. Both strains were grown on glucose or glycerol using ammonia as nitrogen source in batch cultivations and the transcriptome was analyzed using three biological replicated transcriptome experiments. Two areB gene deletion replicates, one on glucose and one on glycerol were discarded due to bad quality and therefore not included in the analysis. Samples for RNA extraction were collected and further processed for hybridization in custom designed Affymetrix microarrays containing probes for three Aspergillus species including A. niger. Triplicate batch fermentations with the two Aspergillus niger strains used, the wild type A. niger strain ATCC 1015 and the areB complete gene deletion strain were carried out and transcriptome analysis was performed. Biomass from each batch cultivation was harvested in the exponential phase of growth and further processed for RNA extraction and hybridization on Affymetrix microarrays.
Project description:This approach aims at searching unidentified regulatory roles of the AreB transcription factor in the overall carbon metabolism of A. niger. A full areB gene deletion mutant was constructed and characterized in A. niger ATCC 1015. Both strains were grown on glucose or glycerol using ammonia as nitrogen source in batch cultivations and the transcriptome was analyzed using three biological replicated transcriptome experiments. Two areB gene deletion replicates, one on glucose and one on glycerol were discarded due to bad quality and therefore not included in the analysis. Samples for RNA extraction were collected and further processed for hybridization in custom designed Affymetrix microarrays containing probes for three Aspergillus species including A. niger.
Project description:Welan gum is mainly produced by Sphingomonas sp. ATCC 31555 and has broad applications in industry such as that in cement production. Both carbon and nitrogen sources are essential for welan production. However, how nitrogen sources affect the metabolism and gene transcription of welan remains elusive. Here, we used next-generation sequencing RNA-seq to analyze the transcriptome of Sphingomonas sp. ATCC 31555 in the presence of inorganic or organic nitrogen sources. Enriched gene expression and pathway analysis suggest that organic nitrogen sources significantly enhanced the expression of genes in central metabolic pathways of Sphingomonas sp. ATCC 31555 and those critical for welan synthesis compared to that observed using inorganic nitrogen sources. The present study improves our understanding of the molecular mechanism underlying the use of nitrogen in welan synthesis in Sphingomonas sp., as well as provides an important transcriptome resource for Sphingomonas sp. in relation to nitrogen sources. Sphingomonas sp. ATCC 31555 strain (stored in our laboratory) was first seeded in an inoculum medium (20 g/L glucose, 3 g/L yeast extract, 3 g/L malt extract, and 5 g/L fish meal protein peptone, pH 7.0), and then cultured in a fermentation medium containing 40 g/L sucrose, 4.0 g/L nitrogen source, 0.6 g/L KH2PO4, and 0.2 g/L MgSO4.7H2O at 37°C. The nitrogen sources used in the present study were as follows: NaNO3 (4.0 g/L) as inorganic nitrogen (IN), beef extract (4.0 g/L) as organic nitrogen (ON), and NaNO3 (1.5 g/L) + beef extract (2.5 g/L) as complex nitrogen (CN). All cultivations were conducted in flasks with constant rotary shaking at 400â??1,000 rpm and 37°C.
Project description:Background: Anaerobic Saccharomyces cerevisiae cultures require glycerol formation to re-oxidize NADH formed in biosynthetic processes. Introduction of the Calvin-cycle enzymes phosphoribulokinase (PRK) and ribulose-1,5-biphosphate carboxylase/oxygenase (RuBisCO) has been shown to couple re-oxidation of biosynthetic NADH to ethanol production and improve ethanol yield on sugar in fast-growing batch cultures. Since growth rates in industrial ethanol-production processes are not constant, performance of engineered strains was studied in slow-growing cultures. Results: In slow-growing anaerobic chemostat cultures (D = 0.05 h-1), an engineered PRK-RuBisCO strain produced 80-fold more acetaldehyde and 30-fold more acetate than a reference strain. This observation suggested an imbalance between in vivo activities of PRK-RuBisCO and formation of NADH in biosynthesis. Lowering the copy number of the RuBisCO-encoding cbbm expression cassette from 15 to 2 reduced acetaldehyde and acetate yields by 67% and 29%, respectively. Additional C-terminal fusion of a 19 amino-acid tag to PRK reduced its protein level by 13-fold while acetaldehyde and acetate production decreased by 94% and 61%, respectively, relative to the 15x cbbm strain. These modifications did not affect glycerol production at 0.05 h-1 but caused a 4.6 fold higher glycerol production per amount of biomass in fast-growing (0.29 h-1) anaerobic batch cultures than observed for the 15x cbbm strain. In another strategy, the promoter of ANB1, whose transcript level positively correlated with growth rate, was used to control PRK synthesis in a 2x cbbm strain. At 0.05 h-1, this strategy reduced acetaldehyde and acetate production by 79% and 40%, respectively, relative to the 15x cbbm strain, without affecting glycerol yield. The maximum growth rate of the resulting strain equalled that of the reference strain, while its glycerol yield was 72% lower. Conclusions: Acetaldehyde and acetate formation by slow-growing cultures of engineered S. cerevisiae strains carrying a PRK-RuBisCO bypass of yeast glycolysis was attributed to an in vivo overcapacity of PRK and RuBisCO. Reducing the capacity of PRK and/or RuBisCO was shown to mitigate this undesirable byproduct formation. Use of a growth-rate-dependent promoter for PRK expression highlighted the potential of modulating gene expression in engineered strains to respond to growth-rate dynamics in industrial batch processes.
Project description:Genome-wide expression analysis of 6 batch cultivations of actinorhodin-producing wild type and recombinant strain of Streptomyces coelicolor
Project description:Transcriptomic analysis was applied to acidogenic, solventogenic and alcohologenic steady-state C. acetobutylicum cells for understanding the regulation of the metabolism of this organism. Used microarray is Clostridium acetobutylicum ATCC824 transcriptional 44k v4, correspondance to GPL10908 Phosphate-limited continuous cultures of C. acetobutylicum were performed under acidogenesis (pH 6.3, 995 mM of carbon from glucose), solventogenesis (pH 4.4, 995 mM of carbon from glucose) and alcohologenesis (pH 6.3, 498 mM of carbon from glucose and 498 mM of carbon from glycerol). Samples were collected for three biological replicates from one control strain and three metabolic mutants, respectively. Total RNA of 36 samples was extracted using an RNeasy Midi kit (Qiagen, Courtaboeuf, France) following the manufacturer's instructions with the supplementation of DNase treatment using RNase-Free DNase Set (Qiagen). Agilent microarray for Clostridium acetobutylicum ATCC 824 (4x44K AMADID 024247) was used. The RNAs were labeled with a Low Input Quick Amp Labeling kit and hybridized following a one-color microarray-based gene expression analysis protocol.
Project description:We report high-throughput RNA sequencing of Δcrp and five adaptively evolved mutants of Δcrp in the batch exponential phase, IG116-Δcrp in batch exponential phase and Δcrp and two adaptively evolved mutants of Δcrp in chemostat cultivations
Project description:NaCl-resistant Saccharomyces cerevisiae mutant was obtained by evolutionary engineering. EMS mutagenized culture was used as the initial population for the selection procedure. Gradually increasing levels of NaCl stress was applied through 40 successive batch cultivations. The reference strain could not grow even at 0.85 M NaCl whereas this mutant was shown to be resistant up to 1.45 M NaCl concentration. Whole-genome microarray analysis was used to identify the NaCl resistance mechanisms by comparing NaCl-resistant mutant strain and the reference strain in the absence of NaCl stress.
