While, many have undergone RT-PCR Test in the last year, but rarely do people know and understand the science behind it! If a housekeeping gene cannot be found whose amplification efficiency is similar to the target, then the standard curve method is preferred RT-PCR reactions on brain, embryo, liver, and spleen total RNA using A primers for clathrin, B primers for clathrin and 18S, or C primers for clathrin, 18S rRNA primers and 18S rRNA Competimers
Molecular Beacons Like TaqMan probes, Molecular Beacons also use FRET to detect and quantitate the synthesized PCR product via a fluor coupled to the 5' end and a quench attached to the 3' end of an oligonucleotide substrate A positive SARS-CoV-2 real-time reverse transcription polymerase chain reaction RT-PCR test is a reliable indicator of infection


Researchers have been able to isolate live virus from hospitalized patients in the first week after symptom onset, with sputum samples producing more live virus than nasopharyngeal swabs.

What is an RT
SARS-CoV-2 viral load in sputum and nasopharyngeal samples varies over time
Therefore, the C t values from PCR reactions run under different conditions or with different reagents cannot be compared directly
How do I interpret RT
Note that without Competimers, 18S cannot be used as an internal control because of its high abundance B
This decouples the fluorescent and quenching dyes and FRET no longer occurs Swab and saliva samples were split between 2 laboratories, which may have led to reporting bias
The RT in the RT-PCR test stands for Reverse Transcription and it an important step in the testing process because of SARS-CoV-2 Virus is RNA based virus They might be infected but we cannot see it yet on RT-PCR

Nucleic Acid Amplification Testing (e.g. RT

Real-time PCR, also called quantitative PCR or qPCR, can provide a simple and elegant method for determining the amount of a target sequence or gene that is present in a sample.

A nasopharyngeal nose or oropharyngeal mouth swab is taken to collect samples for COVID-19 diagnosis
Since RNA codes for protein production in cells, RT-PCR can be used to study the expression of genes in the body
Therefore, the absolute C t value comparison is only meaningful when comparing experiments using the same reaction conditions as defined above
Innovations in NAAT include improved algorithms for handling false negative or false positive results, pooling, faster turnaround times and reductions in the use of materials Thus, for a reliable low copy detection, a large number of replicates are necessary to provide statistical significance and to overcome the Poisson distribution limitation
At some later cycle the amplification rate drops to near zero plateaus , and little more product is made Viral load measurements via RT-PCR

The Basics: RT

Ct was found to be statistically associated with positive culture OR 0.

The PCR product from the endogenous transcript is then compared to the concentration curve created by the synthetic "competitor RNA
How do I interpret RT
They include both DNA deoxyribonucleic acid and RNA ribonucleic acid
Explained: How is COVID
Since the dye binds to double-stranded DNA, there is no need to design a probe for any particular target being analyzed