Table 3.
Primers | PCR master mixed (25 μL total volume) | PCR steps |
---|---|---|
Forward: Se658-F 5′-TTTGATCCTTTGGGTGGTGG-3′ | 2.5 μL of 10× FastStart High Fidelity Reaction buffer with 18 mM MgCl2 (Roche, Mannheim, Germany), 200 μM of dNTP, 0.2 μM of primer (Invitrogen, Carlsbad, CA), 0.625 units of FastStart High Fidelity Enzyme Blend (Roche). | Denatured 94°C 5 mins, 35 cycles at 95°C 30 sec, 59°C for 30 sec, 72°C 45 sec, a final extension at 72°C for 10 min. |
Reverse: Se1124-R 5′-ACCACAAACCACGTGTCATG-3′ |