Polymerase Chain Reaction
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PCR (Polymerase Chain Reaction) is the quick and easy method of making unlimited copies of any fragment of DNA. Since it’s first introduction ten years ago, PCR has very quickly become an essential tool for “improving human health and human life (TPCR)”. Medical research and clinical medicine are profiting from PCR mainly in two areas: detection of infectious disease organisms, and detection of variations and mutations in genes, especially human genes. Because PCR can amplify unimaginably tiny amounts of DNA, even that from just one cell, “physicians and researchers can examine a single sperm or track down the elusive source of a puzzling infection” (TPCR)”. These PCR- based analyses are proving to be just as reliable as previous methods-sometimes more so and often much faster and cheaper.
Polymerase chain reaction (PCR) is a technique used “to amplify the number of copies a specific region of DNA (Brown)”, in order to produce enough DNA to be adequately tested. This technique can be used to identify with a very high-probability, disease-causing viruses, bacteria, a deceased person, a criminal suspect, and also in the event of an outbreak, “Real-Time PCR can effectively monitor the success of clean-up efforts (RAL,Inc)”.
In order to use PCR, one must already know the exact sequence of a gene or the sequence of interest that lie on both ends of the DNA. While similarity among genes of organisms exists, there will always be genes whose DNA sequences differ from each other. By identifying the genes that are different and unique, one can use this information to identify an organism.
DNA is a double-stranded, “consisting of two such nucleotide chains that wind around each other in the famous shape known as the double helix (TPCR)”. DNA consist of Adenine, Thymine, Cytosine, and Guanine components which can be arrange to generate a “sentence” of a gene sequence which can consists of either a few or thousands of letters long.
To get this copying process started, a template molecule of the DNA or RNA you want to copy is required along with two primer molecules that make up the strands of all genetic materials.
These primer molecules consists of about 20 letters long, which can be linked together in the order desired by a DNA-synthesizer “which add and link one letter at a time (Brown)” to generate the primers needed to start PCR.
There are three major steps in PCR that must be met in order for the process to be successful.
During the first step, the desired DNA to be sequenced must be unwound and separated by heating it between “90 to 96 degrees Celsius (TPCR)”. By heating the DNA sample the paired strands separate “becoming individual single strands allowing access for the primers (Brown)”.
The second step of PCR is annealing, where “large excess of primers relative to the amount of DNA being amplified (Brown)” is added to the sample.
During this second stage, the primers and samples are exposed to 54 degrees Celsius and are “joggling around, constantly forming ionic bonds between the standard primer and the single stranded template (POTP)”. Afterwards, the sample mixture is then cooled down, allowing for the double strands to form again.
The third step in PCR is the attachment of an enzyme “DNA-synthesis known as a polymerase (TPCR)” to the opposite ends of the DNA strands. These enzymes are