Uracil-DNA Glycosylase, heat-labile

Uracil-DNA Glycosylase excises uracil from dU-containing DNA by cleaving the N-glycosidic bond between the uracil base and the sugar backbone. This cleavage generates alkali sensitive apyrimidinic sites that are blocked from replication by DNA polymerase or prevented from becoming a hybridization site.

Double and single-stranded dU-containing DNA are substrates for Uracil-DNA Glycosylase whereas RNA and normal dT-containing DNA are not.

Uracil-DNA Glycosylase can also be used to increase the cloning efficiency of PCR products having dU-containing primers incorporated into them as well as increasing the efficiency of site-directed mutagenesis.

The Uracil-DNA Glycosylase derived from Gadus morhua has all the attributes of the enzyme derived from E. coli with the added benefit of being heat-labile. It is completely and irreversibly inactivated after 10 minute incubation at 50°C.

Purity:
The enzyme is chromatographically purified and tested for contaminating endonucleases and exonucleases.

Storage Buffer:
50 mM Tris-HCl (pH 7.5), 100 mM NaCl, 0.5 mM EDTA, 1 mM DTT, 0.1% Triton X-100, 50% glycerol.

Assay Conditions:
The reaction mixture contains 70 mM Tris-HCl, pH 8.0, 10 mM NaCl, 1 mM EDTA, 100 μg/mL BSA, ³H-dUTP labeled DNA, and Uracil-DNA Glycosylase. Incubation is at 37°C for 1 hour.

Unit Definition:
One unit is the amount of enzyme required to liberate 1 nmol uracil from dU-containing DNA in one hour at 37°C.

Concentration:
1 unit/μL

Applications:
  1.Study of DNA repair and mutation detection.
  2.Increase cloning efficiency of PCR products.
  3.Increase the efficiency of site-directed mutagenesis.
  4.Study of protein-DNA interactions.

Source:
Recombinant from Gadus morhua (cod liver)