Polymerase Chain Reaction

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molecular biology assays
based on the detection of specfic nucleic acid sequences in microorganisms or particular cells
polymerase chain reaction
most widely used amplification technique
enables researchers to produce millions of copies of a specific DNA sequence in approximately two hours.
in nature, msot organisms copy their DNA in the same way. The PCR mimics this process, only it does it in a test tube.
principles of PCR
Amplifies low levels of specific DNA sequences in a sample to higher quantitities.
Relies on the ability of DNA-copying enzymes to remain stable at high temperatures
two short DNA primers are used as templates that follow te enzymatic process
Enzymatic process is carried out in a 3-step cycle in the same vial, but at different temperatures.
DNA denaturation
first step
separation of the double DNA strands into two single strands
accomplished by heating the vial to 90-75 degrees centrigrade (about 165 degrees Fahrenheit) for 30 seconds
Primer Annealing
second step
the primers cannto bind to the DNA strangs at such a high temperature, so the vial is cooled to 55 degrees C.
The enzyme polymerase synthesizes new complementary strands by the extension of primers
a complete copy of the templates is made
clinical applications
detection of gene mutations that signify early development of cancer and wide variety of diseases.
PCR :
Human Papilloma Virus (HPV), Coronary Artery Disease
RT-PCR
Human Immunodeficiency Virus, Hepatitis C virus
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