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In research or diagnosis DNA amplification can be conducted through methods such as: Polymerase chain reaction, an easy, cheap, and reliable way to repeatedly replicate a focused segment of DNA by polymerizing nucleotides, a concept which is applicable to numerous fields in modern biology and related sciences.
A strip of eight PCR tubes, each containing a 100 μL reaction mixture 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.
The polymerase chain reaction is the most widely used method for in vitro DNA amplification for purposes of molecular biology and biomedical research. [1] This process involves the separation of the double-stranded DNA in high heat into single strands (the denaturation step, typically achieved at 95–97 °C), annealing of the primers to the single stranded DNA (the annealing step) and copying ...
During the exponential amplification phase, the quantity of the target DNA template (amplicon) doubles every cycle. For example, a DNA sample whose C q precedes that of another sample by 3 cycles contained 2 3 = 8 times more template. However, the efficiency of amplification is often variable among primers and templates.
Loop-mediated isothermal amplification (LAMP) primers [1] Loop-mediated isothermal amplification (LAMP) product [1]. In LAMP, the target sequence is amplified at a constant temperature of 60–65 °C (140–149 °F) using either two or three sets of primers and a polymerase like Bst Klenow fragment with high strand displacement activity in addition to a replication activity.
Similarly, qPCR utilizes fluorescence; however, it measures the intensity of fluorescence at specific times (generally after every amplification cycle) to determine the relative amount of target molecule (DNA), but cannot specify the exact amount without constructing a standard curve using different amounts of a defined standard. It gives the ...
Multiplex-PCR consists of multiple primer sets within a single PCR mixture to produce amplicons of varying sizes that are specific to different DNA sequences. By targeting multiple sequences at once, additional information may be gained from a single test run that otherwise would require several times the reagents and more time to perform.
The PCR method is extremely sensitive, requiring only a few DNA molecules in a single reaction for amplification across several orders of magnitude. Therefore, adequate measures to avoid contamination from any DNA present in the lab environment (bacteria, viruses, or human sources) are required.