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Assessing the effects of transcription factor knockouts on growth in high-density fed-batch cultures

ABSTRACT: Assessing the effects of transcription factor knockouts on growth in high-density fed-batch cultures

PROVIDER: PRJNA915398 | ENA |

REPOSITORIES: ENA

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Assessing the effects of transcription factor knockouts on growth in high-density fed-batch cultures

Project description:We used RNA-seq to investigate the effects of single transcription factor knockouts on growth of E. coli BW25113 in fed-batch cultures at multiple timepoints. Cells were grown in an ambr automated bioreactor and maintained at 30 degrees and pH 6.5 in Medium C.

2022-12-30 | GSE221706 | GEO

Analyzing the metabolic stress response of recombinant Escherichia coli cultures expressing human interferon beta in high cell density fed batch cultures using time course transcriptomic data

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Analyzing the metabolic stress response of recombinant Escherichia coli cultures expressing human interferon beta in high cell density fed batch cultures using time course transcriptomic data

Project description:Over expression of recombinant proteins is known to cause a metabolic burden to the host cells which leads to down regulation of both growth rates and protein expression. Most studies in this regard have been conducted in low density shake flask cultures which does not capture the essential features of an industrial scale bioprocess. In the present work we studied the transcriptomic profiling at different specific growth rates while expressing the recombinant human interferon beta in fed batch cultures with complex media. These conditions mimicked the industrial fermentations for recombinant proteins. To determine the transcriptomic response of E.coli BL21(DE3) to recombinant protein production at three different specific growth rates, samples were collected for four time points (0h, 2h, 4h, 6h) post induction for the fed batch cultivations at specific growth rates 0.2h-1, 0.3h-1, 0.4h-1 respectively and induced at an optical density of 30. 0h (uninduced) sample was taken as control conditions for every run.

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Global transcriptome response of recombinant protein xylanase producing E. coli in high cell density fed batch culture

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Yarrowia lipolytica fed-batch biolipid production.

Project description:Time-course transcriptomic profilling of the oleaginous yeast Yarrowia lipolytica, during a controlled fed-batch. A nitrogen limitation was applied during the course of the fed-batch to initiate de novo biolipid synthesis.

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Global transcriptome response of recombinant protein xylanase producing E. coli in high cell density fed batch culture

Project description:Strong production of recombinant proteins interfere with cellular processes in many ways. The extent of the bacterial stress response is determined by the specific properties of the recombinant protein, and by the rates of transcription and translation. The consideration of bacterial stress and starvation responses is of crucial importance for the successful establishment of an industrial large scale bioprocess. Stress genes can be used as marker genes in order to monitor the fitness of industrial bacterial hosts during fermentation processes. For this purpose, here in our study we have applied transcriptome analysis for the description of general and specific stress and starvation responses of Escherichia coli. Producing recombinant protein (Xylanase) in high cell density fed batch culture. Cells were grown in batch mode in TB media till an OD600 of 15. The specific growth rate of 0.3 h-1 was maintained by fed batch started at OD600 of 15. Cells were induced at an OD600 of 30 using 1 mM IPTG and the growth and product profile was monitored. Samples at different time points (2hrs, 4hrs, and 6hrs) were taken and frozen in liquid nitrogen quickly for transcriptome analysis.

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Batch cultures magnetite, polyaniline

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Transcriptomic analysis of Pichia pastoris (Komagataella phaffii) GS115 during heterologous protein production using a high-cell-density fed-batch cultivation strategy

Project description:We investigated gene expression during the bench-scale batch fermentation phase, glycerol feed phase, glycerol-methanol mixture feed (GM) phase, and at different time points following methanol induction using RNA-Seq. We report that the addition of the GM phase may help to alleviate the adverse effects of methanol addition (alone) on P. pastoris cells. Secondly, enhanced upregulation of the mitogen-activated protein kinase (MAPK) signaling pathway was observed in P. pastoris following methanol induction. The MAPK signaling pathway may be related to P. pastoris cell growth, and may regulate the AOX1 promoter via regulatory factors activated by methanol-mediated stimulation. Thirdly, the unfolded protein response (UPR) and ER-associated degradation (ERAD) pathways were not significantly upregulated during the methanol induction period. These results imply that the presence of unfolded or misfolded phytase protein did not represent a serious problem in our study. Finally, the upregulation of the autophagy pathway during the methanol induction phase may be related to the degradation of damaged peroxisomes but not to the production of phytase. This work describes the metabolic characteristics of P. pastoris during heterologous protein production under high-cell-density fed-batch cultivation. We believe that the results of this study will aid further in-depth studies of P. pastoris protein expression, regulation, and secretory mechanisms.

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Assessing the effects of four two-component system response regulator knockouts in E. coli

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High-cell-density fed-batch cultivations of Vibrio natriegens.

Project description:ObjectivesWith generation times of less than 10 min under optimal conditions, the halophilic Vibrio natriegens is the fastest growing non-pathogenic bacterium isolated so far. The availability of the full genome and genetic engineering tools and its ability to utilize a wide range of carbon sources make V. natriegens an attractive host for biotechnological production processes. However, high-cell-density cultivations, which are desired at industrial-scale have not been described so far.ResultsIn this study we report fed-batch cultivations of V. natriegens in deep-well plates and lab-scale bioreactor cultivations at different temperatures in mineral salt medium (MSM). Upon switching from exponential glucose to constant glucose-feeding cell death was induced. Initial NaCl concentrations of 15-18 g L-1 and a temperature reduction from 37 to 30 °C had a positive effect on cell growth. The maximal growth rate in MSM with glucose was 1.36 h-1 with a specific oxygen uptake rate of 22 mmol gCDW-1 h-1. High biomass yields of up to 55 g L-1 after only 12 h were reached.ConclusionsThe shown fed-batch strategies demonstrate the potential of V. natriegens as a strong producer in industrial biotechnology.

| S-EPMC8397650 | biostudies-literature

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