| 1. | Cut a 1mM thick slice of mouse tail which has been washed with sterile water to remove surface contamination. |
| 2. | Place this section of tail in a 0.5ml PCR tube. |
| 3. | Add 50µl 1XTE. |
| 4. | Add 20µl GeneReleaser®. |
| 5. | Add 2µl PK @ 14-15 mg/ml and mix well. |
| 6. | Digest on thermal cycler 1-3 hours @ 55°C. |
| 7. | Vortex to resuspend. |
| 8. | Heat inactivate 95°C for 10 minutes. (This is critical - DO NOT SHORTEN TIME OR LOWER TEMPERATURE - IF PK IS NOT COMPLETELY INACTIVATED IT WILL DIGEST TAQ.) |
| 9. | Centrifuge 5 minutes @ 10,000xg. |
| 10. | Carefully transfer supernatant to fresh 1.5ml sterile screw cap tube or other suitable storage container. |
| 11. | Use 1µl, 2.5µl, and 5µl of supernatant as template for 3-100µl reactions. This is a rangefinding step. Thereafter use whichever performed best. |
