Quantitative PCR Update

Courtesy of Applied Biosystems  THE TAQMAN PROCEDURE: Applied Biosystems' TaqMan procedure relies on the 5'-to-3' exonuclease activity of Taq polymerase. The TaqMan probe bears a fluoro-phore at the 5' end and a quencher on the 3' end, rendering the molecule non-fluorescent. During amplification, the probe binds to the template between the two PCR primers. When the polymerase encounters the probe, it starts chewing away at the end, releasing the fluorophore into solution, where it is free

Wendy Gloffke
Apr 20, 2003
Courtesy of Applied Biosystems
 THE TAQMAN PROCEDURE: Applied Biosystems' TaqMan procedure relies on the 5'-to-3' exonuclease activity of Taq polymerase. The TaqMan probe bears a fluoro-phore at the 5' end and a quencher on the 3' end, rendering the molecule non-fluorescent. During amplification, the probe binds to the template between the two PCR primers. When the polymerase encounters the probe, it starts chewing away at the end, releasing the fluorophore into solution, where it is free to fluoresce.

In the nearly two decades since its discovery, the polymerase chain reaction (PCR) and its derivative techniques have revolutionized molecular biology. The exponential nature of the technique allows one, in theory, to calculate the amount of starting material from the amount of product at any point in the reaction. In practice, however, reaction conditions can interfere with the exponential amplification and affect product concentration. Standard PCR works best, therefore, as a qualitative...

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