28] Removal of DNA contamination in polymerase chain reaction reagents by ultraviolet irradiation

Abstract

This chapter discusses the removal of DNA contamination in polymerase chain reaction (PCR) reagents by ultraviolet (UV) irradiation. The experiments described demonstrate the efficacy of UV irradiation as a decontamination agent. The method involves mixing the reagents, irradiating with UV for the desired time, adding template DNA, and amplifying with PCR. Although the experiments focus on PCR, UV irradiation can inactivate undesired nucleic acids in other protocols, such as in vitro transcription and certain alternative amplification systems. Two major practical conclusions emerge from these studies. First, it is advantageous not to amplify small segments of DNA whenever feasible because these are likely to be poorly inactivated by UV if these segments become contaminants. Second, UV inactivation is much less effective in eliminating dried DNA, suggesting that the decontamination of laboratory equipment may require the aid of a photosensitizer or even a completely different approach.