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Uracil-DNA Glycosylase, heat-labile

from marine bacterium BMTU 3346, expressed in E. coli

For further processing only.

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Order information
Uracil-DNA Glycosylase, heat-labile material number and pack size:
Material Number Pack Size
11780565103 custom fill
Will be supplied as "Uracil-DNA Glycosylase, heatlab". Unit of measure is "kU".
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Overview
Uracil-DNA Glycosylase (UNG) with an increased heat intolerance is the enzyme of choice for prevention of PCR carryover contamination.
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Application
Use Uracil-DNA Glycosylase, heat-labile, (UNG) for prevention of carryover contamination with DNA amplification products in PCR. Always use dUTP containing PCR mixtures to enable decontamination by UNG treatment. In contrast to the UNG variant from E.coli, this heat-labile enzyme is completely inactivated in the initial heat denaturation step of a common PCR protocol and the formed PCR product will not be degraded.
Note: For high sensitive real-time PCR, specially optimized LightCycler® Uracil-DNA Glycosylase is recommended.
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Specification
Uracil-DNA Glycosylase hydrolyzes uracil-glycosidic bonds in DNA, creating abasic sites where the DNA is cleaved by heat, alkali, or endonuclease treatment. This heat-labile enzyme is easily inactivated by heat denaturation.
Specificity: Hydrolyzes uracil-glycosidic bonds in single- and double-stranded DNA; no activity on dU-free natural DNA and RNA.
Incubation: +15 to +25°C for 10 minutes are recommended for treatment of PCR mixtures; at higher temperatures the enzyme stability decreases.
Half life at +40°C: About 2 minutes
Heat inactivation: +95°C for 2 minutes are sufficient for inactivation
pH optimum: 8.3-8.9
Inhibition: Activity does not depend on metal ions; no inhibition in presence of EDTA or other chelating reagents.
Appearance: Clear, colorless solution
Storage buffer: Tris/HCl, 20 mmol/L; KCl, 100 mmol/L; DTT, 1 mmol/L; EDTA, 0.1 mmol/L; glycerol, 50% (v/v); Nonidet P40, 0.5% (v/v); Tween 20, 0.5% (v/v); pH 8.0 at +4°C
pH value: 8.0±0.1
Volume activity: ≥1 U/μL
Unit definition: One unit Uracil-DNA Glycosylase, heat-labile, is defined as the amount of enzyme required to completely degrade 1 μg purified single-stranded uracil-containing DNA (bacteriophage M13, grown in E.coli CJ236 dut-ung-) at +37°C within 60 minutes. For comparison, one Lindahl unit is comparable to 520,000 units based on our unit definition. One Lindahl unit is defined as the amount of enzyme required to release 1 mol uracil at +37°C in 1 minute.
Unspecific endonucleases (MWM III DNA and M13mp9 ssDNA): Not detectable in up to 20 U after 16 hours incubation at +37°C.
Nicking activity (pBR322 DNA): Not detectable in up to 20 U after 16 hours incubation at +37°C.
Ribonucleases (MS2 RNA): Not detectable in up to 10 U after 4 hours incubation at +37°C.
Exonucleases (3H-DNA): Not detectable in up to 3 U after 4 hours incubation at +37°C.
Function test, DNA decontamination (complete elimination of 10,000 copies of uracil-containing template DNA in a PCR assay): Corresponds to specification
Animal-derived additives: None
Stability: At -15 to -25°C within specification range for 18 months.