HK™-UNG Thermolabile Uracil N-Glycosylase

Applications

• DNA repair studies.

HK™-UNG is a Uracil N-Glycosylase that can be easily heat-inactivated. Uracil N-Glycosylase (also known as uracil-DNA glycosylase) hydrolyzes the N-glycosidic bond between the deoxyribose sugar and uracil in DNA containing deoxyuridine in place of thymidine. HK-UNG is active on both ssDNA and dsDNA that contain uracil, but has no activity on RNA or 2´-deoxyuridine- 5´-monophosphate. HK-UNG is ideal for studying repair of abasic sites in double-stranded DNA.

Unit Definition: One unit of HK-UNG catalyzes the release of 1 nmol of uracil from uracil-containing DNA in 1 hour at 37°C under standard assay conditions.

Note: EPICENTRE's unit is 5- to 20-fold more active than a unit as defined by other suppliers. Therefore, Dilution Buffer is provided for applications requiring lower concentrations of enzyme.

Dilution and Storage Buffer: 50% glycerol containing 50 mM Tris-HCl (pH 7.5), 100 mM NaCl, 0.1 mM EDTA, 1 mM DTT, and 0.1% Triton® X-100.

Quality Control: HK™-UNG is free of detectable exo- and endonuclease and RNase activities.

Figure 1. Brief heat treatment of HK™-UNG eliminates its ability to digest dU-containing DNA. One unit of each uracil N-glycosylase was incubated for 10 minutes at the indicated temperature in 20 µl of a Tris-acetate buffer (pH 8.3). Following heat treatment, ~300 ng of a dU-containing, 450-bp DNA fragment were added and the samples were incubated at 37°C for 30 minutes. Five µl of a stop buffer were added and the samples were separated in a 1.4% agarose gel. The gel was stained with ethidium bromide and the DNA was visualized under UV illumination. The presence of the 450-bp band in Lanes 5 and 7 demonstrates that HK-UNG is inactivated at 70°C and 95°C.