Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Determination of variability of fermentor-grown Aspergillus niger

ABSTRACT: Knowledge of the biological and technical variation for fermentor-grown Aspergillus niger cultures is needed to design DNA microarray experiments properly. We cultured A. niger in batch-operated fermentor vessels and induced with D-xylose. Transcript profiles were followed in detail by qPCR for 8 genes. A variance components analysis was performed on these data to determine the origin and magnitude of variation within each process step for this experiment. 6 Fermentor cultures were selected to determine technical and biological variation for all 14554 ORFs present on this array type. Keywords: Validation of microarrays; variation analysis; experimental design For 5 weeks, 4 batch-fermentation per week were run in which A. niger was grown on 100 mM sorbitol. At 14 hours after oxygen supply had switched from headspace to sparger-inlet each fermentor was induced with either 0.1 mM D-xylose or 0.1 mM sorbitol. Samples were harvested just before induction and 1 hour after induction. Per week, 3 fermentors were induced with D-xylose and 1 fermentor was induced with sorbitol. Six samples were selected to be put on microarrays based on their biomass density, time distribution, magnitude of xylose-induced genes as measured by qPCR, fermentor vessel number.

ORGANISM(S): Aspergillus niger

SUBMITTER: Leo De Graaff

PROVIDER: E-GEOD-11405 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Analysis of variance components reveals the contribution of sample processing to transcript variation.

van der Veen Douwe D Oliveira José Miguel JM van den Berg Willy A M WA de Graaff Leo H LH

Applied and environmental microbiology 20090220 8

The proper design of DNA microarray experiments requires knowledge of biological and technical variation of the studied biological model. For the filamentous fungus Aspergillus niger, a fast, quantitative real-time PCR (qPCR)-based hierarchical experimental design was used to determine this variation. Analysis of variance components determined the contribution of each processing step to total variation: 68% is due to differences in day-to-day handling and processing, while the fermentor vessel, ...[more]

PMID: 19233957

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Project description:The industrially important fungus Aspergillus niger feeds naturally on decomposing plant material, for which it is equipped with a range of enzyme systems. A significant proportion of plant material are lipids that might be available either as for energy storage or as membrane building blocks. With 63 potential lipase-encoding genes in its genome, A. niger has the tools to degrade these extracellular lipids. In contrast to polysaccharide-degrading enzyme networks not much is known about the signalling and regulatory processes that control lipase expression and activity in fungi both under laboratory and natural occurring conditions. A pulse of 1 mM of various oils was applied to four bioreactor-grown A. niger cultures to examine (i) whether A. niger responds at the level of gene transcription, (ii) at what time point this effect is detected most accurately, and (iii) whether differences between the response towards oils are observed. The triglyceride olive oil induces genes encoding peroxins and enzymes of fatty acid metabolism. A complex oil mixture extracted from wheat gluten, which is enriched for digalactosyl-diglycerides, induces genes encoding peroxins as well as enzymes of fatty acid metabolism, but with different expression profile when compared to olive oil. Pure digalactosyldiglyceride, a proxy for plant membrane lipids, does not trigger a transcriptional response. Keywords: time course; induction experiment In one week, 4 fermentor cultures were run in 2.2-liter batch fermentors in which A. niger was grown on 100 mM sorbitol. At 14 hours after oxygen supply had switched from headspace to sparger-inlet each fermentor was induced with 22 mL medium which contained 100 mM (final concentration in fermentor: 1 mM) of either olive oil, a complex oil mixture extracted from wheat gluten, pure wheat digalactosyldiglycerides, or was induced with a solution of minimal medium containing only 0.2% (in fermentor, final concentration 0.002%) triton X-100 which served as emulsifier agent. For each fermentor vessel, a sample of 10 mL was taken prior to induction (T=0), or 30 minutes, 1 hour, or 2 hours after induction. Every sample was hybridized onto a single microarray, yielding in total 16 DNA microarrays.

Dataset Information

Determination of variability of fermentor-grown Aspergillus niger

Publications

Analysis of variance components reveals the contribution of sample processing to transcript variation.

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