Gene Expression
While identifying the sequence of a genome offers insight into what its genes can do, its expression profile provides vital information on what it is doing at any given moment. Gene expression mechanisms in a cell act as both an ‘on/off switch’ to direct which genes are expressed, and as a ‘volume control’ in regulating gene expression. Measurement of coding and non-coding RNA in specific gene transcription processes enables discovery of new regulatory pathways, validation of drug targets, and diagnosis of disease.
Gene expression analysis via quantitative reverse transcription PCR (RT-qPCR) is a key enabling technology of the post-genome era, and is now an established tool for this purpose. RT-qPCR is widely used as a diagnostic technique for the detection of RNA viruses and particular genetic abnormalities leading to disease. In research settings, RT-qPCR is mainly used to provide quantitative measurements of expressed mRNA. This is useful in determining how the expression of a particular gene changes over time, as in the case of cell response to drug treatment. Although the assay method of choice when a gene or pathway is already known, RT-qPCR has limited use for target discovery in that only a relatively small number of genes can be surveyed at a time. Large-scale expression profiling experiments are required to identify target candidates.
Due to their ability to simultaneously monitor expression in thousands of genes at once in a single experiment, microarrays are now used routinely to conduct profiling studies on a genome-wide scale. Microarrays are used to investigate the molecular basis of genetic disease, define groups of marker genes classifying a specific condition, and to screen for subtle response changes in therapeutic treatment. They may also be used to assay gene expression within a single sample or for comparative expression between two samples such as healthy and diseased tissue. However, because of the semi-quantitative nature of microarrays, candidate genes identified during discovery are often validated using RT-qPCR.
Caliper products enable gene expression laboratories to automate sample preparation for RT-qPCR and microarray analysis while also ensuring the quality of results through RNA integrity analysis. The Sciclone® G3 Workstation and Zephyr® Molecular Biology Workstation are ideal for RNA extraction, purification, and RT-qPCR reaction setup. In addition, Sciclone and Zephyr can be integrated with the Roche LightCycler and Twister II Plate Server for a complete, fully automated solution for RT-qPCR applications. The LabChip® GX is quickly becoming the gold standard for quantitation and integrity analysis of mRNA and cRNA prior RT-qPCR and microarray experiments, which is essential for high-quality downstream results. Analysis can be performed in as little as 2 µL of sample, and the platform’s powerful software suite provides further assurance of RNA integrity and consistency by providing quality metrics as well as comparative analysis to reference samples.
