Transcription profiling of E. coli HMS174(DE3) to analyze the cellular response to high level export of recombinant protein to the periplasm
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ABSTRACT: This transcription profiling time course experiment was conducted in order to analyze the cellular response of E. coli HMS174(DE3)-pET30a-sSpAD-GFPmut3.1-Strep to recombinant protein export to the periplasm. Three biological replicates were generated by using a carbon limited exponential fed-batch cultivation.
Project description:This transcription profiling time course experiment was conducted in order to analyze the cellular response of the plasmid-free expression system E.coli BL21(DE3)::TN7<T7 -SOD> to high level expression of recombinant human super-oxide-dismutase (SOD).Three biological replicates were generated by using a carbon limited exponential fed-batch cultivation similar to industrial setups for large scale production. For induction of the system a single pulse of isopropyl-beta-D-galactoside (IPTG) yielding in a fully induced system is applied one doubling past feed start.
Project description:The objective of the current study was to evaluate the performance of frequently used E. coli K-12 strains HMS174 and RV308 in the context of industrial cultivations. To meet this objective, we carried out batch cultivations with high-glucose media in computer-controlled bioreactors and applied transcriptome analysis to interpret phenotypic attributes at the molecular level. The resulting enhancement of knowledge about host physiology is expected to contribute to the acceleration of improvements and optimizations in related biotech processes.
Project description:The objective of the current study was to evaluate the performance of frequently used E. coli BL21 in the context of industrial cultivations. To meet this objective, we carried out batch cultivations with high-glucose media in computer-controlled bioreactors and applied transcriptome analysis to interpret phenotypic attributes at the molecular level. The resulting enhancement of knowledge about host physiology is expected to contribute to the acceleration of improvements and optimizations in related biotech processes.
Project description:This transcription profiling time course experiment was conducted in order to analyze the cellular response of E. coli HMS174(DE3)-pET30a-GFPmut3.1-Strep to high level cytoplasmic protein expression. Three biological replicates were generated by using a carbon limited exponential fed-batch cultivation.
Project description:This transcription profiling time course experiment was conducted in order to analyze the cellular response of the T7 RNA polymerase based E. coli expression system HMS174(DE3)(pET30aNproGFP) to a limited induction with isopropyl-beta-D-galactoside (IPTG). The used cell material was produced under well controlled and defined conditions in an exponential carbon limited fed-batch cultivation at a growth rate of 0.1 per hour. One doubling past feed start induction was started by continuously feeding limiting amounts of inducer gaining in a constant IPTG to cell dry weight ratio of 1µmol/g. Due to the low induction level a physiologically tolerable recombinant gene expression rate was generated and production period was significantly elongated. Non induced cells sampled at the time point of induction were used as reference and compared to cells subsequently sampled during the production period (sampling frequency 2 hours).
Project description:This transcription profiling time course experiment was conducted in order to analyze the cellular response of E. coli BL21(DE3) to high level expression of recombinant human super-oxide-dismutase (SOD) on the transcriptional level. Three biological replicates were generated by using a carbon limited exponential fed-batch cultivation similar to industrial setups for large scale production. For induction of the system a single pulse of isopropyl-beta-D-galactoside (IPTG) yielding in a fully induced system is applied one doubling past feed start. In order to achieve a proper timeresolution of cellular responses sampling starts with a high frequency for the first hours past induction and decreases in the course of the experiment.
Project description:Analysis of the transcriptomes of cell grown at different pH levels pH6.0, pH7.5, pH9.0; Cells were grown in a fermenter at different pH values to an oD600 between 4.5 to 11.5
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