Reaction Mix:
| Tail DNA | 1.0ul |
| 10x buffer | 2.0ul |
| Primer 1 (20uM) | 0.4ul |
| Primer 2 (20uM) | 0.4ul |
| Primer 3 (20uM) | 0.4ul |
| milliQ | 14.8ul |
| Taq (1/25) | 1.0ul |
PCR Primers:
| Primer 1 | 5′ TTC TCA GAT GGT ACC GCT CC 3′ |
| Primer 2 | 5′ GGT GCT TTG AGG CCC GGA TC 3′ |
| Primer 3 | 5′ GAA AGC GAA GGA GCA AAG CTG 3′ |
PCR Conditions:
| 1. | 94C | 3:00 |
| 2. | 94C | 1:00 |
| 3. | 60C | 1:30 |
| 4. | 72C | 2:00 |
| 5. | Goto 2 x30 cycles | |
| 6. | 72C | 5:00 |
| 7. | 4C | 0:00 |
| END | ||
10X PCR Buffer:
| 500 mM KCl |
| 100mM Tris pH 8.3 |
| 15 mM MgCl2 |
| 1 mg/ml BSA |
| 2 mM dNTPs |
| dH20 |
(store at -20C)
Expected Bands:
| Wt= ~550 bp |
| Mut= ~450 bp |