| Step | Temperature (°C) | Time (min) |
|---|---|---|
| 1. Denaturation | 95 | 5 |
| 2. Incubation | 60 | 25 |
| 3. Denaturation | 95 | 5 |
| 4. Incubation | 60 | 85 |
| 5. Denaturation | 95 | 5 |
| 6. Incubation | 60 | 175 |
| Repeat step 1–6 one more time (two cycles of step 1–6 in total) | - | - |
| 7. Hold | 20 | Indefinite |
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