From 2012.igem.org
Restriction Digest
We prepared Master Mixes containing the following volumes,
| EcoR1 + Spe1 Master Mix |
| Substrate | Volume (µL) |
| 10x Buffer | 20 |
| EcoR1 | 2.5 |
| Spe1 | 2.5 |
| Water | 75 |
| Total | 100 |
|
| Xba1 + Pst Master Mix |
| Substrate | Volume (µL) |
| 10x Buffer | 20 |
| Xba1 | 2.5 |
| Pst | 2.5 |
| Water | 75 |
| Total | 100 |
|
| EcoR1 + Pst Master Mix |
| Substrate | Volume (µL) |
| 10x Buffer | 20 |
| EcoR1 | 2.5 |
| Pst | 2.5 |
| Water | 75 |
| Total | 100 |
|
Protocol
- Mix 10 µL of the desired DNA/plasmid and 10 µL of the appropriate Master Mix in a PCR tube.
- Using a thermocycler, incubate the mixture at 37°C for 30 minutes and then deactivate the enzymes by heating to 80°C for 20 minutes.
- Store in a -20°C freezer until DNA required
"
