Multiplex Analyses Using Real-Time Quantitative PCR
Quantitative polymerase chain reaction (qPCR) is a routinely used method for the detection and quantitation of gene expression in real time. Multiplex qPCR requires the use of probe-based assays, in which each probe is labeled with a unique fluorescent dye, resulting in different observed colors for each assay. The signal from each dye is used to quantitate the amount of each target separately in the same tube or well. The availability to multiplex therefore allows the measurement of the expression levels of several targets or genes of interest quickly. Here, we describe a method using the SensiFAST and SensiFAST One-Step probe kits which allows simultaneous real-time quantitation of up to 5 amplicons.
Key wordsqPCR Fluorescent dyes Taq polymerase Quantitation mRNA cDNA Amplicon Multiplex analysis
- 8.Böttcher R, Henderson DJ, Dulla K, van Strijp D, Waanders LF, Tevz G et al (2015) Human phosphodiesterase 4D7 (PDE4D7) expression is increased in TMPRSS2-ERG-positive primary prostate cancer and independently adds to a reduced risk of post-surgical disease progression. Br J Cancer 113:1502–1511CrossRefPubMedPubMedCentralGoogle Scholar