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

ABSTRACT: 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, cDNA synthesis, and qPCR measurement each contributed equally to the remainder of variation. The global transcriptional response to d-xylose was analyzed using Affymetrix microarrays. Twenty-four statistically differentially expressed genes were identified. These encode enzymes required to degrade and metabolize D-xylose-containing polysaccharides, as well as complementary enzymes required to metabolize complex polymers likely present in the vicinity of D-xylose-containing substrates. These results confirm previous findings that the d-xylose signal is interpreted by the fungus as the availability of a multitude of complex polysaccharides. Measurement of a limited number of transcripts in a defined experimental setup followed by analysis of variance components is a fast and reliable method to determine biological and technical variation present in qPCR and microarray studies. This approach provides important parameters for the experimental design of batch-grown filamentous cultures and facilitates the evaluation and interpretation of microarray data.

SUBMITTER: van der Veen D

PROVIDER: S-EPMC2675206 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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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:BackgroundThe ratings of physician-patient communication are an important indicator of the quality of health care delivery and provide guidance for many important decisions in the health care setting and in health research. But there is no gold standard to assess physician-patient communication. Thus, depending on the specific measurement condition, multiple sources of variance may contribute to the total score variance of ratings of physician-patient communication. This may systematically impair the validity of conclusions drawn from rating data.ObjectiveTo examine the extent to which different measurement conditions and rater perspectives, respectively contribute to the variance of physician-patient communication ratings.MethodsVariance components of ratings of physician-patient communication gained from 32 general practitioners and 252 patients from 25 family practices in Germany were analyzed using generalizability theory. The communication dimensions "shared decision making", "effective and open communication" and "satisfaction" were considered.ResultsPhysician-patient communication ratings most substantially reflect unique rater-perspective and communication dimension combinations (32.7% interaction effect). The ratings also represented unique physician and rater-perspective combinations (16.3% interaction effect). However, physicians' communication behavior and the observed communication dimensions revealed only a low extent of score variance (1% physician effect; 3.7% communication dimension effect). Approximately half of the variance remained unexplained (46.2% three-way interaction, confounded with error).ConclusionThe ratings of physician-patient communication minimally reflect physician communication skills in general. Instead, these ratings exhibit primarily differences among physicians and patients in their tendency to perceive shared decision making and effective and open communication and to be satisfied with communication, regardless of the communication behavior of physicians. Rater training and assessing low inferential ratings of physician-patient communication dimensions should be considered when subjective aspects of rater perspectives are not of interest.

Dataset Information

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

Publications

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

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