Gene Expression Profiling: qPCR Toolkit for Quality Control
Poster Apr 05, 2012
Švec D., Jacobsson J., Sjöback R., Kubista M.
Gene expression profiling is an exceedingly important tool in functional genomics research and in drug discovery. The general trend in the field is towards higher throughput of less complex samples. Nowadays studies are appearing where expression of tens of genes is measured in hundreds of individual single cells, to be able to characterize cell subtypes and its roles. To identify subpopulations of astrocytes, to define them in molecular terms, and to determine the biological progress of astrocyte maturation on single cell level we developed and optimized protocol for high-throughput gene expression profiling using qPCR together with a system of quality control: ValidPrimeTM - control for genomic DNA background, Interplate calibrator - tool, which is used to compensate for inter-run variation in qPCR and Universal Spike (RNA/DNA), which can be used to detect presence of inhibition in samples of any living species or monitor losses and yield of isolation, or check the integrity of mRNA during sample processing, storage and transporation.
When there is a need to quickly analyze samples using a number of different PCR assays, it is likely that optimal conditions for each assay will not be the same. First, different assays often will require different annealing temperatures for their primers. In addition, amplicons may be designed to be of different lengths and therefore require varying durations of the extension step.READ MORE