UDG/UNG Enzymes
UDG(uracil DNA glycosylase) can catalyze the hydrolysis of the N-glycosidic link between the uracil base and the sugar-phosphate backbone in ssDNA and dsDNA. It can easily control aerosol pollution and is suitable for common molecular biology systems such as PCR, qPCR, RT-qPCR and LAMP.
Specifications
|
Expression Host |
Recombinant E. coliwith uracil DNA glycosylase gene |
|
Molecular Weight |
24. 8kDa |
|
Purity |
≥95% (SDS-PAGE) |
|
Heat Inactivation |
95℃, 5~10 min |
|
Unit Definition |
One unit (U)is defined as the amount of enzyme that required to catalyze the hydrolysis of l μg dU-containing dsDNA in 30 minutes at 25℃. |
Storage
The product should be stored at-25℃~-15°C for two years.
Instructions
1.Preparation of the PCR reaction mixture according to following system
|
Components |
Volume (μL) |
Final concentration |
|
10×PCR Buffer (Mg²+Plus) |
5 |
1× |
|
25 mmol/LMgCl |
3 |
1.5 mmol/L |
|
dUTP(10 mmol/L) |
3 |
0.6 mmol/L |
|
dCTP/dGTP/dATP/dTTP(10mmol/Leach) |
1 |
0.2 mmol/Leach |
|
Template DNA |
X |
- |
|
Primer1 (10μmol/L) |
2 |
0.4 μmol/L |
|
Primer 2 (10μmol/L) |
2 |
0.4 μmol/L |
|
Taq DNA Polymerase (5 U/μL) |
0. 5 |
0. 05 U/μL |
|
Uracil DNA Glycosylase (UDG/UNG), 1 U/μL |
1 |
1 U/μL |
|
ddH₂O |
Up to 50 |
- |
Note: According to the experimental requirements, the final concentration of dUTP can be adjusted between 0.2-0.6 mmol/L, and 0.2 mmol/L dTTP can be added selectively.
2.Amplification procedure
|
Cycle step |
Temperature |
Time |
Cycles |
|
dU-containing template degradation |
25℃ |
10mins |
1 |
|
UDG activation, template initial denaturation |
95℃ |
5~10mins |
1 |
|
Denaturation |
95℃ |
10 sec |
30-35 |
|
Annealing |
60℃ |
20 sec |
|
|
Extension |
72℃ |
30sec/kb |
|
|
Final extension |
72℃ |
5 min |
1 |
Note: The reaction time at 25°C can be adjusted within 5-10mins according to the experimental requirements.
Notes
1.UDG is active in most PCR reaction buffers.
2.Enzymes should be stored in an icebox or on an ice bath when used, and should be stored at-20°C immediately after use.
3.Please wear the necessary PPE, such lab coat and gloves, to ensure your health and safety!
