I noticed we've been using a PCR system in our one-step RT-PCRs that incorporates dUTPs and uracil-DNA glycosylase (UDG). The UDG is not the cod-derived version. Our reverse transcription (RT'n) is carried out at 50°C for 1 hour, which means the UDG is active during this step.

My understanding is that the produced dU-containing cDNA by RT'n is a target of UDG for degradation.

The thing is, 'it has appeared' as though this has not significantly affected our ability to obtain positive results. However, in the context of diagnostics, I am now curious if we have been unknowingly obtaining false-negatives.

We work with crude plant total nucleic acid extracts and test for RNA-virus infections. Is our ability to produce PCR-positive results entirely dependent on virus titer--that is, is it possible for UDG degradation of virus cDNA to exceed RT'ase production of virus cDNA given a low virus titer? Is it possible for this to no longer be the case given a high virus titer?

Anecdotal or conceptual responses are equally appreciated!