Editing Polymerase chain reaction (section)

{{short description|Laboratory technique to multiply a DNA sample for study}}
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[[Image:PCR tubes.png|thumb|A strip of eight PCR tubes, each containing a 100&nbsp;μL reaction mixture]]
[[Image:PCR masina kasutamine.jpg|thumb|Placing a strip of eight PCR tubes into a [[thermal cycler]]]]

The '''polymerase chain reaction''' ('''PCR''') is a method widely used to make millions to billions of copies of a specific [[DNA]] sample rapidly, allowing scientists to amplify a very small sample of DNA (or a part of it) sufficiently to enable detailed study. PCR was invented in 1983 by American [[biochemist]] [[Kary Mullis]] at [[Cetus Corporation]]. Mullis and biochemist [[Michael Smith (chemist)|Michael Smith]], who had developed other essential ways of manipulating DNA, were jointly awarded the [[Nobel Prize in Chemistry]] in 1993.<ref name="NobelPrize" />

PCR is fundamental to many of the procedures used in [[genetic testing]] and research, including analysis of [[Ancient DNA|ancient samples of DNA]] and identification of infectious agents. Using PCR, copies of very small amounts of [[DNA sequences]] are exponentially amplified in a series of cycles of temperature changes. PCR is now a common and often indispensable technique used in [[medical laboratory]] research for a broad variety of applications including [[biomedical research]] and [[forensic science]].<ref name="Saiki1">{{cite journal | vauthors = Saiki RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N | title = Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia | journal = Science | volume = 230 | issue = 4732 | pages = 1350–54 | date = December 1985 | pmid = 2999980 | doi = 10.1126/science.2999980 | bibcode = 1985Sci...230.1350S }}</ref><ref name="Saiki2">{{cite journal | vauthors = Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA | title = Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase | journal = Science | volume = 239 | issue = 4839 | pages = 487–91 | date = January 1988 | pmid = 2448875 | doi = 10.1126/science.239.4839.487 | bibcode = 1988Sci...239..487S }}</ref>

The majority of PCR methods rely on [[Thermal cycler|thermal cycling]]. Thermal cycling exposes reagents to repeated cycles of heating and cooling to permit different temperature-dependent reactions—specifically, [[DNA melting#Denaturation|DNA melting]] and [[enzyme]]-driven [[DNA replication]]. PCR employs two main reagents—[[Primer (molecular biology)|primers]] (which are short single strand DNA fragments known as [[oligonucleotide]]s that are a [[Complementary DNA|complementary]] sequence to the target DNA region) and a [[thermostable DNA polymerase]]. In the first step of PCR, the two strands of the DNA double helix are physically separated at a high temperature in a process called [[Denaturation (biochemistry)#Nucleic acid denaturation|nucleic acid denaturation]]. In the second step, the temperature is lowered and the primers bind to the complementary sequences of DNA. The two DNA strands then become [[DNA#Polymerases|templates]] for DNA polymerase to [[enzyme|enzymatically]] assemble a new DNA strand from free [[nucleotide]]s, the building blocks of DNA. As PCR progresses, the DNA generated is itself used as a template for replication, setting in motion a [[chain reaction]] in which the original DNA template is [[Exponential growth|exponentially]] amplified.

Almost all PCR applications employ a [[Thermostable DNA polymerase|heat-stable DNA polymerase]], such as [[Taq polymerase|''Taq'' polymerase]], an enzyme originally isolated from the [[thermophile|thermophilic]] bacterium ''[[Thermus aquaticus]]''. If the polymerase used was heat-susceptible, it would denature under the high temperatures of the denaturation step. Before the use of ''Taq'' polymerase, DNA polymerase had to be manually added every cycle, which was a tedious and costly process.<ref>{{cite journal| doi=10.1525/abt.2012.74.4.9| title=Determining Annealing Temperatures for Polymerase Chain Reaction| journal=The American Biology Teacher| volume=74| issue=4| pages=256–60| year=2012| last1=Enners| first1=Edward| last2=Porta| first2=Angela R.| s2cid=86708426}}</ref>

Applications of the technique include [[DNA cloning]] for [[DNA sequencing|sequencing]], gene cloning and manipulation, gene mutagenesis; construction of DNA-based [[phylogeny|phylogenies]], or functional analysis of [[gene]]s; [[medical diagnosis|diagnosis]] and [[monitoring (medicine)|monitoring]] of [[genetic disorder]]s; amplification of ancient DNA;<ref name="Ninfa-2009"/> analysis of genetic fingerprints for [[DNA profiling]] (for example, in [[forensic science]] and [[parentage testing]]); and detection of [[pathogen]]s in [[nucleic acid test]]s for the diagnosis of [[infectious disease]]s.