Table 1 PCR mix and condition for HCMV molecular detection and genotyping by Semi-nested PCR
First round PCR mixture | Second round PCR mixture | Amplification | Temperature | Time | ||
|---|---|---|---|---|---|---|
Component | Volume | Component | Volume | Pre-Denaturation | 95◦C | 5' |
DDW | 28 μl | DDW | 33 μl | Denaturation | 94◦C | 1' |
Buffer | 5 μl | Buffer | 5 μl | Annealing | 55◦C | 1' |
Mgcl2 | 2 μl | Mgcl2 | 2 μl | Extension | 72◦C | 1' |
dNTP | 1.5 μl | dNTP | 1.5 μl | Final extention | 72◦C | 8' |
Forward Primer | 1.5 μl | Forward Primer | 1.5 μl | Hold | 4◦C | - |
Reverse 1 Primer | 1.5 μl | Reverse 2 primer | 1.5 μl | |||
Taq DNA polymerase | 0.5 μl | Taq DNA polymerase | 0.5 μl | |||
Total | 50 μl | Total | 50 μl | |||