Reverse+transcriptase+PCR+(RT-PCR)

Reverse transcriptase PCR is a technique in which products of reverse transcription are amplified by a polymerase chain reaction. Reverse transcription is the process by which fragments of DNA (cDNA) are produced from an RNA template by reverse transcriptase enzymes. The cDNA fragments are then amplified by specific primers under specific PCR conditions. The PCR products are finally run on an agarose gel to visualize the PCR products.



PCR is a highly-sensitive technique as it has a low RNA copy number. It is used in the diagnosis of genetic diseases and semi-quantitative RT-PCR is used to insert eukaryotic DNA into prokaryotic DNA (Jack Vanden Heuvel, Department of Veterinary Science and Molecular Toxicology program, 1998). It is possible to obtain exact information from the DNA due to the high specificity of the method. PCR was designed by an oligonucleotide chemist, Kary Mullis on his way to a weekend in the California Mountains. He realized that just like nuclear fission, where one reaction led to another, polymerase in PCR had the same effect.  RT-PCR was used to discover that chronic intestinal inflammation in Crohn’s disease patients is characterized by an increase in Th1 like cytokines (A., 1995). This procedure was also used for the rapid detection of cases with the novel H1N1 virus thereby enabling better pandemic preparedness (Leo, 2009). RT-PCR was also used to detect suitable RNA targets in normal and cancerous human solid tissues (Latham, 2008). Here, miRNA expression levels were normalized.

__Bibliography__  A., N. B. (1995). Altered Thl/Th2 cytokine profiles in the intestinal mucosa of patients with inflammatory bowel disease as assessed by quantitative reversed transcribed polymerase chain reaction (RT-PCR). //Blackwell Science//, 128-135.   Jack Vanden Heuvel, Department of Vetinary Science and Molecular Toxicology program. (1998). The Basics of PCR. //PCR Protocols in Molecular Toxicology//, 2-4.  Latham.G.J, P. a. (2008). Normalization of microRNA expression levels in quantitative RT-PCR Assays Identifying Suitable RNA Targets in Normal and Cancerous Human Solid Tissues. //RNA//, 844-852.

Leo.L, P. C. (2009). Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays. //Clinical Chemistry//, 1555-1558. 