URACIL-DNA GLYCOSYLASE

General description

Uracil-DNA Glycosylase (UNG) contains the highly active recombinant form of the equally named enzyme found in prokaryotes and eukaryotes. It hydrolyzes uracil-glycosidic bonds in single- or double-stranded DNA, excising uracil and creating alkali-sensitive abasic sites in the DNA. These abasic sites can be hydrolyzed by endonuclease, heat, or alkali treatment. Depending on how the DNA is prepared, Uracil-DNA Glycosylase can be used to achieve general, site-specific, or strand-specific U-DNA cleavage.

Specificity

• Uracil-DNA glycosylase hydrolyzes uracil-glycosidic bonds at U-DNA sites in single- and doublestranded DNA, excising uracil and creating alkali sensitive abasic sites in the DNA.
• The enzyme is more active on single-stranded DNA than on double-standed DNA.
• Activity was also observed on small U-DNA oligonucleotides and on dUMP (Duncan, unpublished observations).
• Uracil-DNA glycosylase is inactive on RNA and native, uracil-free DNA.

Heat inactivation: 95 °C for 10 min
Uracil-DNA glycosylase remains partially active (<10%) after an incubation period of 30 minutes at 95 °C.

Application

Uracil-DNA Glycosylase can be used to cleave DNA at any site where a deoxyuridylate residue has been incorporated. U-DNA can be prepared by in vitro methods like PCR. General, site-specific, or strand-specific cleavage can be achieved with uracil-DNA glycosylase, depending on how the U-DNA is prepared. Uracil-DNA Glycosylase can therefore help you to:

• Prevent carryover contamination in PCR
• Increase the efficiency of site-directed mutagenesis procedures
• Label oligonucleotide probes

Quality

Carryover prevention activity is assayed by adding approximately 10⁵dU that contains templates prior to the amplification reaction. After UNG treatment, no amplification products could be detected. The enzyme does not contain any contaminating exo- or endonucleases and is tested for the absence of RNases.

Unit Definition

One unit is defined as the amount of Uracil-DNA Glycosylase necessary to completely degrade 1 μg purified single-stranded uracil containing DNA (bacteriophage M13, grown in E.coli CJ 236 dut-ung-) at +37 °C in 60 minutes.
One Lindahl unit is defined as the amount of enzyme necessary to release of 1 μmol uracil at +37 °C in 1 minute. One Lindahl unit is comparable to 520,000 U based on our unit definition.

Volume Activity: 1 U/μl

Physical form

The enzyme is supplied as 1U/μl solution in storage buffer.

Other Notes

For life science research only. Not for use in diagnostic procedures.