Polymerase chain reaction

PCRPCR amplificationpolymerase chain reaction (PCR)amplificationDNA amplificationapplicationsgene amplificationamplifiedamplifyerror-prone PCR
Polymerase chain reaction (PCR) is a method widely used in molecular biology to make several copies of a specific DNA segment.wikipedia
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Taq polymerase

Taq'' polymeraseTaq DNA polymeraseTaq
Almost all PCR applications employ a heat-stable DNA polymerase, such as Taq polymerase, an enzyme originally isolated from the thermophilic bacterium Thermus aquaticus. A basic PCR set-up requires several components and reagents, including a DNA template that contains the DNA target region to amplify; a DNA polymerase; an enzyme that polymerizes new DNA strands; heat-resistant Taq polymerase is especially common, as it is more likely to remain intact during the high-temperature DNA denaturation process; two DNA primers that are complementary to the 3' (three prime) ends of each of the sense and anti-sense strands of the DNA target (DNA polymerase can only bind to and elongate from a double-stranded region of DNA; without primers there is no double-stranded initiation site at which the polymerase can bind); specific primers that are complementary to the DNA target region are selected beforehand, and are often custom-made in a laboratory or purchased from commercial biochemical suppliers; deoxynucleoside triphosphates, or dNTPs (sometimes called "deoxynucleotide triphosphates"; nucleotides containing triphosphate groups), the building blocks from which the DNA polymerase synthesizes a new DNA strand; a buffer solution providing a suitable chemical environment for optimum activity and stability of the DNA polymerase; bivalent cations, typically magnesium (Mg) or manganese (Mn) ions; Mg 2+ is the most common, but Mn 2+ can be used for PCR-mediated DNA mutagenesis, as a higher Mn 2+ concentration increases the error rate during DNA synthesis;
It is frequently used in the polymerase chain reaction (PCR), a method for greatly amplifying the quantity of short segments of DNA.

Thermal cycler

thermal cyclersthermal cyclingthermocycler
The vast majority of PCR methods rely on thermal cycling.
The thermal cycler (also known as a thermocycler, PCR machine or DNA amplifier) is a laboratory apparatus most commonly used to amplify segments of DNA via the polymerase chain reaction (PCR).

Oligonucleotide

oligonucleotidesoligoantisense oligonucleotide
PCR employs two main reagents – primers (which are short single strand DNA fragments known as oligonucleotides that are a complementary sequence to the target DNA region) and a DNA polymerase.
Commonly made in the laboratory by solid-phase chemical synthesis, these small bits of nucleic acids can be manufactured as single-stranded molecules with any user-specified sequence, and so are vital for artificial gene synthesis, polymerase chain reaction (PCR), DNA sequencing, library construction and as molecular probes.

Thermus aquaticus

TaqThermophilus aquaticusThermus aquaticus'' Vn 4-311
Almost all PCR applications employ a heat-stable DNA polymerase, such as Taq polymerase, an enzyme originally isolated from the thermophilic bacterium Thermus aquaticus.
It is the source of the heat-resistant enzyme Taq DNA polymerase, one of the most important enzymes in molecular biology because of its use in the polymerase chain reaction (PCR) DNA amplification technique.

Primer (molecular biology)

primerprimersRNA primer
PCR employs two main reagents – primers (which are short single strand DNA fragments known as oligonucleotides that are a complementary sequence to the target DNA region) and a DNA polymerase. A basic PCR set-up requires several components and reagents, including a DNA template that contains the DNA target region to amplify; a DNA polymerase; an enzyme that polymerizes new DNA strands; heat-resistant Taq polymerase is especially common, as it is more likely to remain intact during the high-temperature DNA denaturation process; two DNA primers that are complementary to the 3' (three prime) ends of each of the sense and anti-sense strands of the DNA target (DNA polymerase can only bind to and elongate from a double-stranded region of DNA; without primers there is no double-stranded initiation site at which the polymerase can bind); specific primers that are complementary to the DNA target region are selected beforehand, and are often custom-made in a laboratory or purchased from commercial biochemical suppliers; deoxynucleoside triphosphates, or dNTPs (sometimes called "deoxynucleotide triphosphates"; nucleotides containing triphosphate groups), the building blocks from which the DNA polymerase synthesizes a new DNA strand; a buffer solution providing a suitable chemical environment for optimum activity and stability of the DNA polymerase; bivalent cations, typically magnesium (Mg) or manganese (Mn) ions; Mg 2+ is the most common, but Mn 2+ can be used for PCR-mediated DNA mutagenesis, as a higher Mn 2+ concentration increases the error rate during DNA synthesis;
Living organisms use solely RNA primers, while laboratory techniques in biochemistry and molecular biology that require in vitro DNA synthesis (such as DNA sequencing and polymerase chain reaction) usually use DNA primers, since they are more temperature stable.

Molecular cloning

cloneclonedrecombinant DNA technology
Applications of the technique include DNA cloning for sequencing, gene cloning and manipulation, gene mutagenesis; construction of DNA-based phylogenies, or functional analysis of genes; diagnosis and monitoring of hereditary diseases; amplification of ancient DNA; analysis of genetic fingerprints for DNA profiling (for example, in forensic science and parentage testing); and detection of pathogens in nucleic acid tests for the diagnosis of infectious diseases.
Molecular cloning is similar to polymerase chain reaction (PCR) in that it permits the replication of DNA sequence.

Molecular biology

molecular biologistmolecularmolecular microbiology
Polymerase chain reaction (PCR) is a method widely used in molecular biology to make several copies of a specific DNA segment.
In this technique, DNA coding for a protein of interest is cloned using polymerase chain reaction (PCR), and/or restriction enzymes into a plasmid (expression vector).

Thermophile

thermophilicthermophilesthermophilous
Almost all PCR applications employ a heat-stable DNA polymerase, such as Taq polymerase, an enzyme originally isolated from the thermophilic bacterium Thermus aquaticus.
Some of these enzymes are used in molecular biology, for example the taq polymerase used in PCR.

Amplicon

ampliconsamplicon sequencingbands
To check whether the PCR successfully generated the anticipated DNA target region (also sometimes referred to as the amplimer or amplicon), agarose gel electrophoresis may be employed for size separation of the PCR products.
It can be formed artificially, using various methods including polymerase chain reactions (PCR) or ligase chain reactions (LCR), or naturally through gene duplication.

In silico PCR

Electronic PCR
Computer simulations of theoretical PCR results (Electronic PCR) may be performed to assist in primer design.
In silico PCR refers to computational tools used to calculate theoretical polymerase chain reaction (PCR) results using a given set of primers (probes) to amplify DNA sequences from a sequenced genome or transcriptome.

Chain reaction

chain reactionschemical chain reactionchain
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 exponentially amplified.

Gene duplication

duplicationduplicationsamplification
Using PCR, copies of DNA sequences are exponentially amplified to generate thousands to millions of more copies of that particular DNA segment.
Genetic amplification can occur artificially, as with the use of the polymerase chain reaction technique to amplify short strands of DNA in vitro using enzymes, or it can occur naturally, as described above.

Nucleic acid sequence

DNA sequencesnucleotide sequencegenetic information
Using PCR, copies of DNA sequences are exponentially amplified to generate thousands to millions of more copies of that particular DNA segment.
This is overcome by polymerase chain reaction (PCR) amplification.

DNA sequencing

DNA sequencesequencesequencing
Applications of the technique include DNA cloning for sequencing, gene cloning and manipulation, gene mutagenesis; construction of DNA-based phylogenies, or functional analysis of genes; diagnosis and monitoring of hereditary diseases; amplification of ancient DNA; analysis of genetic fingerprints for DNA profiling (for example, in forensic science and parentage testing); and detection of pathogens in nucleic acid tests for the diagnosis of infectious diseases.
The DNA sample preparation and random surface-polymerase chain reaction (PCR) arraying methods described in this patent, coupled to Roger Tsien et al.'s "base-by-base" sequencing method, is now implemented in Illumina's Hi-Seq genome sequencers.

Real-time polymerase chain reaction

quantitative PCRqPCRreal-time PCR
Quantitative PCR or Real Time PCR (qPCR, not to be confused with RT-PCR) methods allow the estimation of the amount of a given sequence present in a sample—a technique often applied to quantitatively determine levels of gene expression.
A real-time polymerase chain reaction (real-time PCR), also known as quantitative polymerase chain reaction (qPCR), is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR).

Molecular-weight size marker

molecular weight size markerDNA ladderDNA marker
The size(s) of PCR products is determined by comparison with a DNA ladder, a molecular weight marker which contains DNA fragments of known size run on the gel alongside the PCR products.
This strategy involves the use of Polymerase Chain Reaction (PCR).

Agarose gel electrophoresis

agarose gelagaroseelectrophoresis
To check whether the PCR successfully generated the anticipated DNA target region (also sometimes referred to as the amplimer or amplicon), agarose gel electrophoresis may be employed for size separation of the PCR products.
The lower sulphate content of low EEO agarose, particularly low-melting point (LMP) agarose, is also beneficial in cases where the DNA extracted from gel is to be used for further manipulation as the presence of contaminating sulphates may affect some subsequent procedures, such as ligation and PCR.

Ancient DNA

aDNAextracted from fossilsancient
PCR may also be used in the analysis of ancient DNA that is tens of thousands of years old.
However, with the development of the Polymerase Chain Reaction (PCR) in the late 1980s, the field began to progress rapidly.

Nucleic acid test

nucleic acid amplification testNAATNucleic acid amplification tests
Applications of the technique include DNA cloning for sequencing, gene cloning and manipulation, gene mutagenesis; construction of DNA-based phylogenies, or functional analysis of genes; diagnosis and monitoring of hereditary diseases; amplification of ancient DNA; analysis of genetic fingerprints for DNA profiling (for example, in forensic science and parentage testing); and detection of pathogens in nucleic acid tests for the diagnosis of infectious diseases.
There are several ways of amplification including polymerase chain reaction (PCR), strand displacement assay (SDA), or transcription mediated assay (TMA).

Nucleic acid thermodynamics

annealingmelting temperatureDNA melting
Thermal cycling exposes reactants to repeated cycles of heating and cooling to permit different temperature-dependent reactions – specifically, DNA melting and enzyme-driven DNA replication.
In the absence of external negative factors, the processes of hybridization and melting may be repeated in succession indefinitely, which lays the ground for polymerase chain reaction.

Base pair

base pairsbpMbp
Most PCR methods amplify DNA fragments of between 0.1 and 10 kilo base pairs (kbp) in length, although some techniques allow for amplification of fragments up to 40 kbp.
GC content and melting temperature must also be taken into account when designing primers for PCR reactions.

Infection

infectious diseaseinfectious diseasesinfections
Applications of the technique include DNA cloning for sequencing, gene cloning and manipulation, gene mutagenesis; construction of DNA-based phylogenies, or functional analysis of genes; diagnosis and monitoring of hereditary diseases; amplification of ancient DNA; analysis of genetic fingerprints for DNA profiling (for example, in forensic science and parentage testing); and detection of pathogens in nucleic acid tests for the diagnosis of infectious diseases.
Technologies based upon the polymerase chain reaction (PCR) method will become nearly ubiquitous gold standards of diagnostics of the near future, for several reasons.

Preimplantation genetic diagnosis

preimplantation genetic screeningpre-implantation genetic diagnosisPGD
It was not until the 1980s that human IVF was fully developed, which coincided with the breakthrough of the highly sensitive polymerase chain reaction (PCR) technology.

Influenza

fluhuman fluthe flu
A type of polymerase chain reaction that detects the virus's RNA is more accurate.

Complementarity (molecular biology)

complementarycomplementaritycomplement
A basic PCR set-up requires several components and reagents, including a DNA template that contains the DNA target region to amplify; a DNA polymerase; an enzyme that polymerizes new DNA strands; heat-resistant Taq polymerase is especially common, as it is more likely to remain intact during the high-temperature DNA denaturation process; two DNA primers that are complementary to the 3' (three prime) ends of each of the sense and anti-sense strands of the DNA target (DNA polymerase can only bind to and elongate from a double-stranded region of DNA; without primers there is no double-stranded initiation site at which the polymerase can bind); specific primers that are complementary to the DNA target region are selected beforehand, and are often custom-made in a laboratory or purchased from commercial biochemical suppliers; deoxynucleoside triphosphates, or dNTPs (sometimes called "deoxynucleotide triphosphates"; nucleotides containing triphosphate groups), the building blocks from which the DNA polymerase synthesizes a new DNA strand; a buffer solution providing a suitable chemical environment for optimum activity and stability of the DNA polymerase; bivalent cations, typically magnesium (Mg) or manganese (Mn) ions; Mg 2+ is the most common, but Mn 2+ can be used for PCR-mediated DNA mutagenesis, as a higher Mn 2+ concentration increases the error rate during DNA synthesis;
This principle is the basis of commonly performed laboratory techniques such as the polymerase chain reaction, PCR.