Competitive RNA Templates for Detection and Quantitation of Growth Factors, Cytokines, Extracellular Matrix Components and Matrix Metalloproteinases by RT-PCR

摘要

Detection of low-abundance mRNAs by reverse transcription-polymerase chain reaction (RT-PCR) has become a standard technique to determine gene expression by tissues and cells in culture. The ability to determine relative or absolute copy number of specific mRNAs has been difficult due to inadequate in ternal standards to control for sample-to-sample variation. The use of a synthetic RNA standard with idenficdt sequences to the PCR primers allows reproducible quantitation between samples and assays.By designing multi-sequence templates, several specific mRNAs can be qitan-tiiated using a single template. Addition of multiple templates to a single RT reaction allows the quantitation of a large number of targets from as little as 4 fjg of total RNA,In this report, we present a series of seven primer/template systems to detect and quantitate 52 specific messages, including 26 growth factors and receptors, 8 extracellular matrix components, 10 matrix-modifying enzymes and their inhibitors and 8 cy-tokines.