Real-Time PCR

     The Polymerase Chain Reaction (PCR) is a revolutionary force in the detection of both RNA and DNA. The advent of Real-Time technologies has added functionality to PCR by allowing investigators to visualize the reaction as it takes place.  In normal PCR reactions it is generally assumed that there is a quantitative relationship between the amount of starting nucleic acid and the amount at the end of any one cycle.  In practice this has proven difficult and different reactions yield varying amounts of PCR product.  Real-Time PCR (RT-PCR) has reduced the variability involved with quantifying DNA and RNA.   This allows for a very sensitive, reproducible and accurate system for investigators to utilize. Theoretically, as little as a single copy of nucleic acid can be amplified and detected utilizing real-time technologies.    As well as using RT-PCR to quantify the amount of DNA, it can also be applied to pathogen identification and detection, viral load, allelic discrimination studies, quality control, assay validation, and genotyping.

o     Application:

· Determining the lower limit of detection of pathogenic DNA.

· Comparison of transcript levels

o     Useful Information:

·  http://www.biosci.ohio-state.edu/~pmgf/procedures_rt-pcr.pdf

·  http://dna-9.int-med.uiowa.edu/realtime.htm#supplinfo

·   http://pga.tigr.org/sop/RT-PCR.pdf

·   http://showelab.wistar.upenn.edu/REAL_TIME_qPCR.pdf

·   http://www.infectio.com/en/recherche/pdf/march-2000.pdf

·   http://www.hmc.psu.edu/core/Genomics_Core/RealTime_PCR/REALTIMEdocs.htm