Department of Biochemistry

Clinical Applications of PCR Related

to Corona Virus

Written by : Rasoul Hashim Hussein

Group : A \ Second stage
ID : 80342
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Introduction
Coronavirus disease (COVID-19) is an infectious disease caused by a newly discovered
coronavirus.

Most people infected with the COVID-19 virus will experience mild to moderate
respiratory illness and recover without requiring special treatment. Older people, and
those with underlying medical problems like cardiovascular disease, diabetes, chronic
respiratory disease, and cancer are more likely to develop serious illness.

The best way to prevent and slow down transmission is be well informed about the
COVID-19 virus, the disease it causes and how it spreads. Protect yourself and others
from infection by washing your hands or using an alcohol based rub frequently and not
touching your face.

The polymerase chain reaction

The polymerase chain reaction (PCR) underlies almost all of modern molecular cloning.
Using PCR, a defined target sequence that occurs once within a DNA of high
complexity and large size-an entire mammalian genome, for example-can be rapidly
and selectively amplified in a quasi-exponential chain reaction that generates millions of
copies. The reaction is simple to set up, cheap, and undemanding, the only requirement
being some knowledge of the nucleotide sequences of the target. In addition to its
simplicity, PCR is robust, speedy, flexible, and sensitive.

PCR involves three basic steps, which occur repeatedly, usually for about 35 to 40
cycles:

1- a denaturation step at 95°C, in which DNA is heated to become single-stranded.

2- an annealing step at a temperature usually between 50° and 60°C, in which an

oligonucleotide primer hybridizes to each of the single DNA strands.

3- an extension step at 72°C, in which an engineered DNA polymerase adds

complementary bases to each of the single DNA strands, creating double-stranded
DNA.

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PCR Test
The PCR test looks for the presence of the covid-19 virus on a sample of tissue from a
nasophayngeal swab or a saliva specimen. Polymerase chain reaction (PCR) tests are
used to detect HIV's genetic material, called RNA. These tests can be used to screen
the donated blood supply and to detect very early infections before antibodies have
been developed. This test may be performed just days or weeks after exposure to HIV.
Although these tests are the most accurate, they are not performed as often as the
other HIV tests because they are expensive and also time- and labor-intensive.

Real time polymerase chain reaction in covid-19

Real time RT–PCR is one of the most widely used laboratory methods for detecting the
COVID-19 virus. While many countries have used real time RT–PCR for diagnosing
other diseases, such as the Ebola virus and Zika virus, many need support in adapting
this method for the COVID-19 virus, as well as in increasing their national testing
capacities

Real time RT–PCR is a nuclear-derived method for detecting the presence of specific
genetic material in any pathogen, including a virus. Originally, the method used
radioactive isotope markers to detect targeted genetic materials, but subsequent
refining has led to the replacement of isotopic labelling with special markers, most
frequently fluorescent dyes. This technique allows scientists to see the results almost
immediately while the process is still ongoing, whereas conventional RT–PCR only
provides results at the end of the process.

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Types of PCR
1- Real-Time PCR (quantitative PCR or qPCR) – in which DNA molecules are
tagged using fluorescent dye, which is used to monitor and quantify PCR
products in real-time
2- Reverse-Transcriptase (RT–PCR) – creates complementary DNA (cDNA) by
reverse transcribing RNA to DNA using reverse transcriptase
3- Multiplex PCR – uses a number of primers to multiply multiple fragments in a
single DNA sample
4- Nested PCR – after the initial 25-35 PCR cycles, an additional PCR is conducted
using new primers “nested” within the original primers, which reduces the risk of
unwanted products
5- Hot Start PCR – in which heat is used to denaturate antibodies that are used to
inactivate Taq polymerase
6- Long-range PCR – longer ranges of DNA are formed by using a mixture of
polymerases
7- Assembly PCR – longer DNA fragments are aplified by using overlapping primers
8- Asymmetric PCR – only one strand of the target DNA is amplified
9- In situ PCR – PCR that takes place in cells, or in fixed tissue on a slide

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References
1. Laboratory testing for Middle East Respiratory Syndrome coronavirus, interim
guidance (revised), January 2019, WHO/MERS/LAB/15.1/Rev1/2019, World Health
Organization, 2018.

2. Managing epidemics, key facts about major deadly diseases. Geneva: World Health
Organization; 2018.

3. WHO Global Influenza Surveillance Network Manual for the laboratory diagnosis
and virological surveillance of influenza, WHO, 2011 .

4. Protocol to investigate non-seasonal influenza and other emerging acute respiratory

diseases. Geneva: World Health Organization; 2018.

5. WHO Recommended Surveillance Standards WHO/CDS/CSR/ISR/99.2 .

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