A real-time polymerase chain reaction (real-time PCR), also termed a quantitative polymerase chain reaction (qPCR), is a molecular biology laboratory technique based on the polymerase chain reaction (PCR). During the PCR (i.e., in real time), it monitors the amplification of a targeted DNA molecule, not at its end, as in conventional PCR. Real-time PCR (quantitative real-time PCR) and semi-quantitative (i.e. above / below a certain amount of DNA molecules) (semi-quantitative real-time PCR) can be used.
Two common methods for detecting PCR products in real-time PCR are non-specific fluorescent dyes intercalating with any double-stranded DNA and sequence-specific DNA samples consisting of oligonucleotides labeled with a fluorescent reporter, which allow detection only after the complementary sequence of the sample has been hybridised. We define a semi-quantitative RT-PCR protocol optimized for extracting RNA from as few as 10,000 cells in our laboratory and measuring the expression rates of multiple target mRNAs from each specimen. This procedure has been optimized on the human erythroleukemia cell line TF-1, but has been used successfully on primary cells and on various cell lines.