The Process of PCR

Authors: Junghyun Kwon (Hannah)

Editors: Vincent Chang and Liane Xu

Artist: Susan Wu

PCR, or polymerase chain reaction, is a method used to prepare billions of copies of specific DNA sequences by amplifying a sample of DNA. As large amounts of DNA samples are needed for molecular and genetic analysis and research, studying DNA is almost impossible without this revolutionary technology of amplification. Today, PCR is used in DNA fingerprinting, diagnosis of genetic disorders, and of course, detection of viruses.

To perform a PCR experiment, a scientist needs primers, nucleotides, Taq polymerase, a DNA template, and a thermocycler. Primers are short fragments of DNA or RNA that bind to a specific complementary region of the DNA template to mark the beginning of DNA synthesis. Nucleotides are building blocks of DNA made up of sugar, a phosphate group, and a nitrogenous base (either A, T, G, or C). Taq polymerase is a thermostable DNA polymerase active even at temperatures up to 95°C, ensuring that multiple cycles of PCR are performed in a single continuous event. DNA template, of course, is the basis from which a new strand is copied. Lastly, a thermocycler raises and lowers the temperature every few minutes to allow for DNA denaturation and synthesis.

There are three distinct steps in the process of PCR: denaturation, annealing, extension, and they are all done inside the thermocycler, each in different temperatures. Firstly, during denaturation, a DNA sample is heated to around 95 degrees, and the two strands are broken apart. Then, at the annealing temperature of around 5 degrees below the primer melting point, the primers attach to the template strand. After that, at 72 degrees, extension temperature, Taq polymerase binds to the primer and starts copying the template DNA by adding the complementary nucleotides. This cycle of denaturation and synthesizing DNA is continuously repeated and by the end of 30 cycles, more than a billion copies of the original segment would be present in the PCR tube.

The results of a PCR reaction can be visualized using gel electrophoresis, a technique in which fragments of DNA are pulled through a gel matrix by an electric current, and it separates DNA fragments according to size.

This amazing technique is frequently used nowadays to test for COVID-19. By copying the sample of DNA from the inside of your nose and creating a billion copies, we can tell whether or not the virus is present.

 

Citations:

Polymerase chain reaction (PCR) fact sheet. Genome.gov. (n.d.).

https://www.genome.gov/about-genomics/fact-sheets/Polymerase-Chain-Reaction-Fact-Sheet

Khan Academy. (n.d.). Polymerase chain reaction (PCR) (article). Khan Academy.

https://www.khanacademy.org/science/ap-biology/gene-expression-and-regulation/biotechnology/a/polymerase-chain-reaction-pcr.

PCR test for COVID-19: What it is, how its done, what the results mean. Cleveland Clinic. (n.d.).

https://my.clevelandclinic.org/health/diagnostics/21462-covid-19-and-pcr-testing.

6 views0 comments