Absolute estimation of initial concentrations of amplicon in a real-time RT-PCR process

Background: Since real time PCR was first developed, several approaches to estimating the initial quantity of template in an RT-PCR reaction have been tried. While initially only the early thermal cycles corresponding to exponential duplication were used, lately there has been an effort to use all of the cycles in a PCR. The efforts have included both fitting empirical sigmoid curves and more elaborate mechanistic models that explore the chemical reactions taking place during each cycle. The more elaborate mechanistic models require many more parameters than can be fit from a single amplification, while the empirical models provide little insight and are difficult to tailor to specific reactants. Results: We directly estimate the initial amount of amplicon using a simplified mechanistic model based on chemical reactions in the annealing step of the PCR. The basic model includes the duplication of DNA with the digestion of Taqman probe and the re-annealing between previously synthesize...

• Type: Journal
• Year: 2007
• Where: BMCBI
• Authors: Marjo V. Smith, Chris R. Miller, Michael Kohn, Nigel J. Walker, Chris J. Portier
• Keywords: BMCBI 2007, Chemical Reactions, Elaborate Mechanistic Models, Taqman Probe