RT-PCR can be performed in either a one-step or a two-step
assay. 

In a one-step assay, reverse transcription and PCR
amplification are consolidated into one reaction. This assay format can provide rapid and reproducible results for high-throughput analysis. The challenge is the difficulty in optimizing the reverse transcription and amplification steps
as they occur simultaneously, which leads to lower target
amplicon generation. 

In the two-step assay, the reaction is done sequentially in separate tubes. This assay format is more sensitive than the one-step assay, but it is more time consuming and requires optimizing additional parameters.

Controls need to be carefully selected to ensure the reliability of the assay and to identify experimental errors.

University of California, Santa Cruz

Smith College

RNA is reverse transcribed into cDNA and amplified using a polymerase chain reaction. Primers target various sections of the virus genetic sequence with the E gene being the highest sensitivity test and being confirmed by the RdRp gene.
Upper and lower respiratory samples are collected. Nasopharyngeal swabs are high priority specimens for SARS-CoV-2 while low priority specimens include: sputum, oropharyngeal swabs, and tracheal aspirates.

RT-PCR

Udugama B, Kadhiresan P, Kozlowski H, Malekjahani A, Osborne M, Li V, Chen H, Mubareka S, Gubbay J, and Chan W

doi: 10.1021/acsnano.0c02624

Diagnosing COVID-19: The Disease and Tools for Detection

False positives can be caused from the cross-reactivity of primers with nucleic acids which happen because contamination of other bacteria and viruses. If reagents of RT-PCR false positives can also occur which is a problem considering the high volume of testing occurirng in laboratories during the pandemic. 
A simpler method of diagnosing COVID-19 where resources are limited would be more beneficial.

Disadvantages of RT-PCR

Advantages: RT-PCR can analyze thousands of samples in a day at a sensitivity of 95%. Positives should generally follow the trend of clinical observations and epidemiological information.

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