Methods in molecular cardiology: quantitative real-time polymerase chain reaction

Changes in mRNA expression levels occur during physiological and pathological processes in the cardiovascular system. An increase in DNA transcription results in increased mRNA levels and will subsequently cause an increase in levels of proteins that regulate processes inside and outside the cell. To determine alterations in mRNA levels, traditional methods such as Northern blotting and ribonuclease protection assay can be used; however, large amounts of RNA are necessary and the methods are very labour intensive. In this review, we focus on one of the newest advancements in polymerase chain reaction (PCR) technology, real-time or quantitative PCR, using small amounts of RNA to determine expression levels. We discuss the technique in general and describe two different approaches.