False Negative Results from Using Common PCR Reagents

Background:
The sensitivity of the PCR reaction makes it ideal for use when identifying potentially novel viral infections in human disease. Unfortunately, this same sensitivity also leaves this popular technique open to potential contamination with previously amplified PCR products, or "carry-over" contamination. PCR product carry-over contamination can be prevented with uracil-DNA-glycosylase (UNG), and it is for this reason that it is commonly included in many commercial PCR master-mixes. While testing the sensitivity of PCR assays to detect murine DNA contamination in human tissue samples, we inadvertently discovered that the use of this common PCR reagent may lead to the production of false-negative PCR results.

Findings:
We show here that contamination with minute quantities of UNG-digested PCR product or any negative control PCR reactions containing primer-dimers regardless of UNG presence can completely block amplification from as much as 60ng of legitimate target DNA.

Conclusions:
The potential for false negative results needs to be considered and carefully controlled in PCR experiments, especially when the target copy number may be low - just as the potential for false positive results already is.
 
The article is published online in BMC Research Notes and is free to access.