Table 3 Optimization of the RAPD-PCR reaction parameters for amplification of genomic DNA of Passiflora foetida
Sr no | PCR parameters | Tested range | Optimum conditions | Inferences |
|---|---|---|---|---|
1 | DNA concentration (ng) | 50, 75, 100, 150, 200 | 100 ng | Less amplification with lower concentration and smear formation at higher concentration |
2 | Primer concentration (μM) | 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5. | 2 μM | Minimum amount produces sufficient amplification |
3 | Dream Taq polymerase (units) | 2 U, 5 U | 2 U | Sufficient for proper amplification. |
4 | Annealing temperature (°C) | 25, 27, 29, 30, 35, 40,44 | 29°C | Lower annealing temperatures show proper annealing and amplification |
5 | No of cycles | 25, 30, 35, 40, 45 | 30 | Higher/lower cycles (from optimum) effects the amplification |