Quantitative “Real-Time” (RT) PCR

          The Polymerase Chain Reaction (PCR) is ordinarily a preparative or qualitative procedure used to produce a large quantity of DNA for analysis. Use of PCR primers tagged with a fluorescent reporter dye allows the quantitative progress of a PCR amplification experiment to be monitored in “real time.” The number of cycles required for the amplification to shift from the initial linear to the exponential phase is an indication of abundance of the original template [left]. The inflection points (slopes) of two reactions with known initial concentrations can be used to draw a standard curve [right], which allows the number of gene copies in the unknown samples to be calculated.

          Biodiversity applications: Use of species-specific PCR primer sequences would allow monitoring of any one species against a diverse background, for example, the relative abundance of Atlantic Cod in the stomach contents of Harp Seals.