Measurement of cardiac gene expression by reverse transcription polymerase chain reaction (RT-PCR).

Two methods applicable to the measurement of cardiac gene expression by reverse transcription polymerase chain reaction (RT-PCR) are presented. Each method describes a specific technique and includes information on how to optimize the experiments and how to trouble-shoot any problems. The first method illustrates how conventional RT-PCR was used to detect expression of a specific gene in rat heart. The second method explains how to investigate the amount of gene expressed under different conditions utilizing the new technique of quantitative real-time RT-PCR. More specifically, this second method was used to investigate the effect of a relevant cardiac insult, ischemia reperfusion, on gene expression, although the technique could be readily adapted to other cardiac insults or to compare different hearts or treatments. The gene investigated in these examples was SNAT3, which is responsible for the cotransport of sodium predominantly with glutamine in exchange for hydrogen ions. The results suggested that a 20-fold increase in SNAT3 expression occurs during ischemia, which reduced to baseline levels on reperfusion.