The use of qRT-PCR and high-throughput transcriptomics for biomarker development

Michael W Pfaffl, Irmgard Riedmaier, Heinrich HD Meyer
Physiology Weihenstephan, TUM, 85354 Freising, Germany

The detection of growth promoting agents is a central topic in human doping surveillance and agricultural meat production. In the focus are the discovery and early verification of the illegal drug application, e.g. hormone cocktails or new xenobiotic growth promoters. In routine screening, the residues of all known growth promoters are detected by immuno based assays or chromatographical methods in combination with mass spectrometry. To overcome the detection by these routine applied screening methods, in the previous years new xenobiotic drugs and application techniques were adapted. To enable a future efficient tracing of misused anabolic substances it is necessary to develop new screening and detection technologies for a broad range of illegal hormones, including newly designed xenobiotic anabolic agents. To achieve this, many innovative technologies have been described for biomarker discovery and validation. This talk provides an overview of the latest transcriptomics based technologies in biomarker discovery after illegal hormone application. These –omics technologies can provide essential information on the mRNA expression of hormone regulated genes across multiple tissues, organs, treatment stages, and experimental trials. It highlights recent gene expression results on mRNA and microRNA level derived from quantitative RT-PCR and next generation sequencing. On the basis of the expression results reliable candidate biomarker were selected. To verify if these gene expression changes could act as first valid biomarkers bio-statistical methods were applied. Principal Components Analysis and Hierarchical Clustering were successfully used to demonstrate the potential of the transcriptomic approach for the development of a new screening system and to introduce new gene expression biomarkers.

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