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In light of recent events, the term PCR is not foreign to many of us. PCR, also known as Polymerase Chain Reaction, is a revolutionary method developed by Kary Mullis in the 1980s. It is based on using the ability of DNA polymerase (an enzyme that creates DNA molecules by assembling nucleotides - which are subunits of DNA) to synthesize new strands of DNA complementary to the offered template strand. At the end of a PCR reaction, we end up with billions of copies of a specific DNA fragment or gene, which enables the visual detection and identification of gene sequences based on size.
All PCR analysis requires the presence of DNA polymerase, nucleotides, primers and template DNA. DNA polymerase is an essential enzyme that joins the individual nucleotides together to form the PCR product. Nucleotides are the four bases found in DNA - adenine, thymine, cytosine and guanine (A, T, C, G). They act as building blocks that DNA polymerase uses to create the resulting PCR product. Primers are short fragments of DNA with a defined sequence that is complementary to the target DNA. They specify the exact section of DNA that needs to amplified by the DNA polymerase
These components are all mixed in a test tube and placed in a thermocycler that allows repeated cycles of DNA amplification. First, the solution is heated to allow the target DNA strands to separate (denaturation). The temperature is then lowered to allow specific primers to bind to target DNA segments (annealing). This is followed by an increase in temperature that causes the DNA polymerase to attach to the primers, adding nucleotides to the developing DNA strand.
The PCR method does have drawbacks, such as the fact that the DNA polymerase used in the PCR, is prone to errors which can cause the fragment generated to be incorrect. However, the likelihood of such errors occuring is very low.
PCR continues to remain unmatched on the basis of importance due to the precision of the technique and the impact it has globally. The COVID-19 RT-PCR Test is a test for the detection of nucleic acid from SARS-CoV-2, Real time RT–PCR is one of the most widely used laboratory methods for detecting the COVID-19 virus, the Ebola virus and even Zika virus.
Written by Elizza Miriam Mathew
References
How is the COVID-19 virus detected using real time RT–PCR? | IAEA. (n.d.). International Atomic Energy Agency. Retrieved September 4, 2023, from https://www.iaea.org/bulletin/how-is-the-covid-19-virus-detected-using-real-time-rt-pcr
Polymerase Chain Reaction (PCR). (2017, November 9). NCBI. Retrieved September 2, 2023, from https://www.ncbi.nlm.nih.gov/probe/docs/techpcr/
Real-Time Polymerase chain reaction trends in COVID-19 patients. (n.d.). NCBI. Retrieved September 2, 2023, from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7794159/
Research Techniques Made Simple: Polymerase Chain Reaction (PCR). (n.d.). NCBI. Retrieved September 2, 2023, from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4102308/
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