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Polymerase Chain Reaction (PCR) is a molecular biological method for amplifying (creating multiple copies of) DNA without using a living organism, such as E. coli or yeast. PCR is commonly used in medical and biological research labs for a variety of tasks, such as the detection of hereditary diseases, the identification of genetic fingerprints, the diagnosis of infectious diseases, the cloning of genes, and paternity testing.1 History
140px Kary Mullis was awarded a Nobel Prize for the invention of PCR
The basic method for performing PCR was invented by Kary Mullis, who was awarded the Nobel Prize in Chemistry in October 1993 for this achievement, only seven years after he first published his ideas. Mullis's idea was to develop a process by which DNA could be artificially multiplied through repeated cycles of duplication driven by an enzymeAn enzyme is a protein, or protein complex, that catalyzes a chemical reaction. Like any catalyst, enzymes work by lowering the activation energy of a reaction, thus allowing the reaction to proceed to its steady state or completion much faster than it ot called DNA polymeraseA DNA polymerase is an enzyme that assists in DNA replication. Such enzymes catalyze the polymerization of deoxyribo nucleotides alongside a DNA strand, which they "read" and use as a template. The newly polymerized molecule is complementary to the templa.
DNA polymerase occurs naturally in living organisms, where it functions to duplicate DNA when cells divide. It works by binding to a single DNA strand and creating the complementary strand. In Mullis's original PCR process, the enzymeAn enzyme is a protein, or protein complex, that catalyzes a chemical reaction. Like any catalyst, enzymes work by lowering the activation energy of a reaction, thus allowing the reaction to proceed to its steady state or completion much faster than it ot was used in vitro (in a controlled environment outside an organism). The double-stranded DNA was separated into two single strands by heating it to 96°C. At this temperature, however, DNA-Polymerase was destroyed so that the enzyme had to be replenished after the heating stage of each cycle. Mullis's original PCR process was very inefficient since it required a great deal of time, vast amounts of DNA-Polymerase, and continual attention throughout the PCR process.
Later, this original PCR process was improved by the use of DNA-Polymerase taken from thermophilic (heat-loving) bacteria that grow in geyserA geyser is a special type of hot spring that erupts periodically, ejecting a column of hot water and steam into the air. The name geyser comes from Geysir the name of the best-known geyser in Iceland; that name, in turn, comes from the word gjosa "to guss at a temperature of over 110°C. The DNA-Polymerase taken from these organisms is thermostable (stable at high temperatures) and, when used in PCR, did not break down when the mixture was heated to separate the DNA strands. Since there was no longer a need to add new DNA-Polymerase for each cycle, the process of copying a given DNA strand could be simplified and automated.
One of the first thermostable DNA-Polymerases was obtained from Thermus aquaticusThermus aquaticus is a species of bacterium that is the source of the enzyme Taq DNA Polymerase, one of the most important enzymes in molecular biology. It is one of several thermophilic bacteria that belong to the Deinococcus-Thermus group. Biology In 19 and called Taq. Taq polymerase is widely used in current PCR practice (May 2004). A disadvantage of Taq is that it sometimes makes mistakes when copying DNA, leading to mutationThis article is about mutation in biology, for other meanings see: mutation (disambiguation). Mutations are permanent, transmissible changes to the genetic material (usually DNA or RNA) of a cell. Mutations can be caused by copying errors in the genetic ms (errors) in the DNA sequence, since it lacks 3'->5' proofreading exonuclease activity. Polymerases such as Pwo or Pfu, obtained from ArchaeaPhylum Crenarchaeota Phylum Euryarchaeota Halobacteria Methanobacteria Methanococci Methanopyri Archeoglobi Thermoplasmata Thermococci Phylum Korarchaeota Phylum Nanoarchaeota The Archaea are a major group of prokaryotes. They were first identified in 197, have proofreading mechanisms (mechanisms that check for errors) and can significantly reduce the number of mutations that occur in the copied DNA sequence. Combinations of both Taq and Pfu are available nowadays that provide both the high fidelity and accurate amplification of DNA
