Notebook > Protocol > PCR with EasyPfu DNA Polymerase
Protocol for PCR with EasyPfu DNA Polymerase
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Preparation for Reaction Mixture
| Reagent | Final concentration | Volume
|
| 10X Taq buffer | 1X | 2 µl
|
| 2.5 mM dNTP mix | 0.25 mM | 2 µl
|
| Forward Primer | 0.2-0.4 µM | 1 µl
|
| Reverse Primer | 0.2-0.4 µM | 1 µl
|
| EasyPfu DNA Polymerase | 25 units | 1 µl
|
| Template DNA | As required | <0.5 µg
|
| ddH2O | to final volume |
|
Cycling Conditions
| Step | Temperature/℃ | Time/min
|
| Initial Denaturation | 94 | 5
|
| Denaturation | 94 | 0.5
|
| Primer Annealing | As required | As required
|
| Extending | 72 | As required(0.5kb/min)
|
| Final Extending | 72 | 10
|
| Number of Cycles | | 32-35
|
| Cooling Down | 4 | 10
|
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