Uracil-DNA glycosylase (UNG), cod is a thermolabile recombinant enzyme produced in E. coli (ung-) using a modified ung gene derived from Atlantic cod. It degrades uracil-containing single- and double-stranded DNA, but not RNA or thymidine-containing DNA, by hydrolysing the N-glycosidic bond between deoxyribose sugar and the base in uracil. This generates alkaline-sensitive apyramidinic sites in the DNA that will be cleaved upon a combination of alkaline conditions and high temperature.
- Complete, irreversible heat-inactivation
- Prevents carryover contamination in PCR, qPCR and qRT-PCR
- Effective with only a 5 minute incubation step
Pre-treatment of samples with UNG prevents PCR carryover contamination in laboratories that substitute dUTP in place of dTTP during all amplification reactions. PCR products containing uracil become substrates for UNG and will be degraded if they are present in subsequent reaction mixtures subjected to UNG treatment. Only DNA templates containing thymidine are not degraded by the treatment and will be amplified.
In contrast to competing Uracil-DNA Glycosylases, VWR Life Science’s arctic-derived UNG has the distinct advantage of having the low inactivation temperature of 45 °C, which enables carryover decontamination of one-step reverse transcription reactions with UNG prior to the cDNA synthesis step. Inactivation is irreversible after heating just 20 minutes at 55 °C or 1 second at 95 °C, allowing for stability of the cDNA that has incorporated dUTP. The irreversible inactivation feature of VWR Life Science UNG also facilitates greater downstream manipulation and stability of newly synthesised dUTP-containing DNA by PCR, allowing for long-term storage, restriction digestion, cloning, sequencing and hybridisation applications. All amplification reactions must use dUTP-containing dNTP mixtures in order for the UNG decontamination method to be effective.
Product is stable at least 2 years when stored frozen (–20 to 0 °C). Product may be stored cold (4 to 8 °C) for up to 6 months.