A MIQE Case Study — Effect of RNA Sample Quality and Reference Gene Stability on Gene Expression Data
Application Note Feb 21, 2012
Real-time quantitative PCR (qPCR) has become the gold standard for validating DNA microarray data and is routinely used to determine gene expression differences between a wide variety of samples. The exquisite sensitivity of the technology permits the detection of a single copy of a target gene in a sample which has led to qPCR now being used in the clinical setting to diagnose infection and disease states. In an effort to standardize the design of the associated experiments, the minimum information for publication of quantitative real-time PCR experiments (MIQE) guidelines were published in 2009. This study shows how qPCR can lead to erroneous conclusions regarding differences in MCM7 gene expression between normal and tumor human breast cancer samples if the key steps set out in the MIQE guidelines are not followed.