Real-time Polymerase Chain Reaction

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Other names: Quantitative PCR, qPCR

The Real-time Polymerase Chain Reaction is an improvised version of the original Polymerase Chain Reaction (PCR) developed by Kary Mullis, who received the Nobel Prize in Chemistry in 1993, and her co-workers during the mid-1980s[1]. They are designed to provide information as fast as the amplification process itself. As the information is showed continuously as the assay proceeds in 'real time', the reaction cycle at which specific PCR products can be identified. The amount of template DNA or RNA present at the beginning of the reaction can then be interpreted[2].

Monitoring of the Real-Time PCR

The product that is formed from the Real-time PCR binds to a required flourescent reporter, a probe that is only developed recently to report the amplification (presence and amount) of PCR product in real-time. The signal is very small and is almost the same as the background during the initial cycles. The amount of product would then increase and the signal would exponentially increase. In the end, the signal would saturate as the reaction does not have enough of the required components (either the primers, the fluorescent reporter or the dNTP[3]) to continue with the reaction[4].

References

  1. Saiki RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anaemia. Science. 1985 Dec 20;230(4732):1350-4.
  2. David Oldach, “Real-time” polymerase chain reaction, Gastroenterology, Volume 116, Issue 3, 1999, Pages 763-765, ISSN 0016-5085
  3. Mikael Kubista, Anders Stalberg, Tzachi Bar. Light-up-probe-based real-time Q-PCR. Geonomics and Proteomics Technologies. 2001; Proceedings volume 4264.
  4. Mikael Kubista, José Manuel Andrade, Martin Bengtsson, Amin Forootan, Jiri Jonák, Kristina Lind, Radek Sindelka, Robert Sjöback, Björn Sjögreen, Linda Strömbom, Anders Ståhlberg, Neven Zoric,The real-time polymerase chain reaction, Molecular Aspects of Medicine,Volume 27, Issues 2–3, 2006, Pages 95-125
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