NaOH treatment to neutralize inhibitors of Taq polymerase

The introduction of polymerase chain reaction (PCR) into the forensic field has greatly extended the ability to analyze DNA from small or degraded sam ples. However, one significant problem with PCR analysis is the sensitivity of Tag Polymerase to inhibitors found in many substrates commonly encounte red with evidentiary materials. We hypothesize that the most problematic of these compounds intercalate into double stranded DNA (dsDNA) and have sign ificantly less affinity for single stranded DNA (ssDNA). This study present s a comprehensive analysis of a novel method for the neutralization of Tag inhibitors by denaturation and washing with NaOH in Microcon-100 filtration units. The data show that DNA recovered following NaOH repurification rout inely amplifies when other inhibitor neutralization techniques are unsucces sful. Genetic profiles have been obtained with both AmpliType PM + DQA1 and D1S80 systems. However, the NaOH protocol is not advised when the quantity of DNA is limited since the treatment results in significant loss of DNA.