Dataset Information

Quantitative PCR deconstruction of discrepancies between results reported by different hybridization platforms.

ABSTRACT: Differences in hybridization platforms used in gene array analysis experiments can lead to significant differences in hybridization results. In this study we used quantitative reverse transcription-polymerase chain reaction (qRT-PCR) to investigate discrepant results between the National Institute of Environmental Health Sciences cDNA and Affymetrix oligo platforms used to evaluate hepatic gene expression changes in rats exposed to methapyrilene. Caldesmon cDNA platform hybridization results showed decreases in gene expression levels for the high-dose methapyrilene 7-day pooled samples compared with their controls. By contrast, the Affymetrix oligonucleotide platform showed increases in expression levels for these samples. Quantitative gene expression measurements provide an explanation for the discrepancies observed for these samples. In the case of caldesmon, there is a 74-base sequence in the cDNA clone that is absent in the Affymetrix sequence. The amplicon based on the cDNA clone shows > 100-fold suppression relative to the day 7 high-dose methapyrilene-pooled control. These data demonstrate the importance of using a "gold standard," such as qRT-PCR to confirm key hybridization results as well as to understand the sources of discrepancies resulting from different hybridization platforms.

SUBMITTER: Goodsaid FM

PROVIDER: S-EPMC1241899 | biostudies-other | 2004 Mar

REPOSITORIES: biostudies-other

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Quantitative PCR deconstruction of discrepancies between results reported by different hybridization platforms.

Goodsaid Federico M FM Smith Roger J RJ Rosenblum I Y IY

Environmental health perspectives 20040301 4

Differences in hybridization platforms used in gene array analysis experiments can lead to significant differences in hybridization results. In this study we used quantitative reverse transcription-polymerase chain reaction (qRT-PCR) to investigate discrepant results between the National Institute of Environmental Health Sciences cDNA and Affymetrix oligo platforms used to evaluate hepatic gene expression changes in rats exposed to methapyrilene. Caldesmon cDNA platform hybridization results sho ...[more]

PMID: 15033595

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Project description:Mixed methods are commonly used in health services research; however, data are not often integrated to explore complementarity of findings. A triangulation protocol is one approach to integrating such data. A retrospective triangulation protocol was carried out on mixed methods data collected as part of a process evaluation of a trial. The multi-country randomised controlled trial found that a web-based training in communication skills (including use of a patient booklet) and the use of a C-reactive protein (CRP) point-of-care test decreased antibiotic prescribing by general practitioners (GPs) for acute cough. The process evaluation investigated GPs' and patients' experiences of taking part in the trial.Three analysts independently compared findings across four data sets: qualitative data collected view semi-structured interviews with (1) 62 patients and (2) 66 GPs and quantitative data collected via questionnaires with (3) 2886 patients and (4) 346 GPs. Pairwise comparisons were made between data sets and were categorised as agreement, partial agreement, dissonance or silence.Three instances of dissonance occurred in 39 independent findings. GPs and patients reported different views on the use of a CRP test. GPs felt that the test was useful in convincing patients to accept a no-antibiotic decision, but patient data suggested that this was unnecessary if a full explanation was given. Whilst qualitative data indicated all patients were generally satisfied with their consultation, quantitative data indicated highest levels of satisfaction for those receiving a detailed explanation from their GP with a booklet giving advice on self-care. Both qualitative and quantitative data sets indicated higher patient enablement for those in the communication groups who had received a booklet.Use of CRP tests does not appear to engage patients or influence illness perceptions and its effect is more centred on changing clinician behaviour. Communication skills and the patient booklet were relevant and useful for all patients and associated with increased patient satisfaction. A triangulation protocol to integrate qualitative and quantitative data can reveal findings that need further interpretation and also highlight areas of dissonance that lead to a deeper insight than separate analyses.

Dataset Information

Quantitative PCR deconstruction of discrepancies between results reported by different hybridization platforms.

Publications

Quantitative PCR deconstruction of discrepancies between results reported by different hybridization platforms.

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