PCR (Polymerase Chain Reaction – Polymerase Chain Reaction) test is a test that is frequently brought up these days due to the coronavirus epidemic. This test, which is used for coronavirus diagnosis, has many uses, especially in the field of health. In this article, the titles such as exactly what the PCR test is, for what purposes it is used, the way the PCR test is applied, and what should be considered when it is applied, how it is used in coronavirus has been examined in detail.
What is the PCR test? What is the PCR test basis?
PCR (Polymerase Chain Reaction – Polymerase Chain Reaction) is a common name given to the reactions applied to enzymatically increase a specific region in DNA. In short, the PCR method is to imitate or repeat DNA replication for a certain number of times. This method can also be described as “in vitro” ie in vitro cloning. PCR test is a molecular recognition technique. PCR test is an advanced and highly sensitive technique among molecular recognition techniques. It can be considered as an advanced technique because it can distinguish even a small number of microbes.
PCR (Polymerase Chain Reaction) technique is based on separating the two strands of DNA by exposing them to high temperature, that is, becoming a single strand, then extending the strand by binding the synthetic oligonucleotides to the target DNA and repeating this cycle for a certain number of times. In this way, this technique multiplies even a small number of microbes and makes them recognizable.
Why is the PCR test done? What are the usage areas of the PCR test?
The PCR test is used in many studies, which are based on diagnosis, and in the determination of unknown features. Examples of the uses of the PCR test in the field of health are given below.
1. Diagnosis of carrier and patient in hereditary diseases: By multiplying gene sequences in the PCR test, defects in these genes can be detected. Being able to diagnose early even in carriers is a great advantage of this test.
2. In prenatal diagnosis: Prenatal abnormalities in the fetus can be detected in this way. These abnormalities are usually caused by genetic disease.
3. Detection of pathogenic organisms in clinical samples: It has a great contribution in distinguishing pathogenic organisms by reproducing even a small number of microbes. It is a fast and sensitive technique in disease diagnosis. In the coronavirus detection, the pathogenic organism can be recognized and diagnosed in this way.
4. In forensic medicine: In this method, DNA analysis of criminal persons can be performed by multiplying the gene sequences performed in the PCR test. Therefore, it has great importance in forensic medicine.
5. In researches related to cancer: Mutations in the genes where the tumor occurs can be detected by the PCR test.
6. In the paternity test,
7. In the creation of probes/cloning/gene expression studies,
8. In DNA sequence analysis, creation of a large number of DNA samples,
9. In the determination of unknown series,
10. In the study of past DNA and enlightenment of evolution,
11. In the development of vaccines in the field of biotechnology,
12. Ensuring the normal birth of the baby by performing pre-implantation genetic tests and then performing implantation in a single cell within Vitro fertilization,
13. Development of techniques used in the diagnosis of genetic inbreeding in many plants and animals such as PCR-derived RAPD, AFLP, SSR, and ISRR,
14. DNA can be used in the investigation of protein interaction (footprinting).
PCR test has many uses in the food industry, as well as its uses in the health field mentioned above.
Polymerase Chain Reaction test use in coronavirus diagnosis
Coronavirus diagnostic tests are of two types as molecular and serological tests. The tests used in the diagnosis of coronavirus can be done with the help of the detection of the genetic material of the virus (it can be DNA or RNA) or the antibodies created by the antigenic material against the virus. The tests used to detect the genetic material of the virus are called molecular tests, while the tests that try to detect the antigen-antibody of the virus are called serological tests. Antibody-antigen tests, that is, serological tests, are tests used to detect the stages of the viral infection that has been or is currently undergoing. A real-time PCR test can be used to confirm serological tests in Coronavirus. The tests to be used can be chosen according to the stage of the disease.
The main thing that the PCR test uses in the diagnosis of coronavirus is to detect the RNA polymerase enzyme gene, that is, its genetic material, which can be found depending on RNA in the virus. This test must be applied by a knowledgeable microbiologist and laboratory. Otherwise, it can lead to very wrong results. As a result of incorrect results, the disease cannot be diagnosed and this can lead to further spread of the virus.
If the results are negative, the possibility of the test being performed incorrectly should be considered. The time the sample was taken and whether the sample was taken correctly is also very important for the test to give an accurate result. The performance of the PCR test varies according to the abundance of RNA in the sample, as it makes the diagnosis from RNA detection. Therefore, the quality of the sample taken is also very important for the efficiency of this test. In addition, improper transfer of the sample taken may cause inaccurate results. Technical errors that are not caused by the person performing the test, such as virus mutation or PCR test inhibition, are among the errors that may occur during the application of the test.
There are two types of applications for PCR at this time. These types are PCR tests that give real-time results and give fast results. While it takes 3-4 hours to get results in real-time PCR tests, this time can be reduced to 1.5 hours in PCR tests that give rapid results. In Turkey, this type of real-time PCR test is used.
In the PCR test applied for coronavirus diagnosis, a sample of the respiratory tract must be taken from the patient, such as a nasopharyngeal swab (swab taken from the nose or throat) or sputum sample from the lower respiratory tract. Different methods can be used to take this sample. Taking samples with throat swabs among these methods is a method that can give correct results for the first week. In the continuation of the disease, that is, when the patient becomes pneumonia, the virus spreads rapidly in the lungs and the number in the throat decreases and disappears, and therefore the sample taken from the throat may not be meaningful for diagnosis. Therefore, since the disease progresses after the first week of the disease, diagnosis can be made through the cough material, sputum.
After the samples are taken at the right times and in the right way, the samples are placed in a liquid viral transport medium that can allow the viral RNA to pass into solution. The RNAs passing from the sample to the solution are then separated from this solution, and this RNA is then amplified to facilitate identification. From the RNA that is replicated, the type of virus that RNA belongs to is determined by a number of tests in the laboratory and thus the diagnosis is made.
PCR; Denaturation performed in the range of 94 ° C-98 ° C, separation of the two strands of DNA exposed to high temperature, annealing between 37 ° C-65 ° C, namely the binding of synthetic oligonucleotides to the target DNA (hybridization) and It consists of 3 stages, namely elongation stages, and these stages are repeated in a certain cycle for a certain number of times.
Besides the coronavirus tests that give direct results, the disadvantage of the PCR test is that it gives results at a later time, but the fact that the PCR method detects even a small number of pathogens indicates that this method is more sensitive than other methods. The fact that this test is more sensitive can be counted among the advantages of this test compared to other tests. The PCR technique is also a very advantageous method as it reduces the need for radioactivity. Radioactive diagnostic methods are generally known for their harm to the body. Because of this feature, the PCR technique has become more prominent and more usable among diagnostic methods.
Coronavirus diagnostic tests applied in the laboratory at the right time and with the right methods are an important part of the process in controlling the epidemic. Therefore, patients who think they have symptoms should apply to the nearest health center.
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