I am from IT/ Engineering background and have some confusion in RT-PCR. So far my understanding we extract RNA from test specimens and transcribed it into complementary DNA (cDNA), and inside PCR DNA double helix is separated. Then primer a short nucleic acid sequence that provides a starting point for DNA synthesis with the target sequence and multiples the target DNA.
Since a higher cp value indicates a negative result but why negative results have cp value? Why it cross the threshold when it does have any target sequence (viral DNA)? Fluorescence is the measure of target DNA right??
In extraction, RNA is extracted does only COVID viral RNA is deducted? What happens to other viral and human RNA?
I have seen result were we have low cp value and test result is negative, the reason for this was because curve do not have Sigmoid function. I need more explanation on it.