What are the critical factors for amplification of GC-rich templates?
What are the critical factors for amplification of GC-rich templates?
1. PCR conditions
-Use higher denaturation temperatures (e.g., 98°C as opposed to 94°C or 95°C) to allow complete denaturation of the template.
-Keep annealing times for GC-rich templates as short as possible.
-Use primers with a higher Tm (>68°C), because annealing can occur at a higher temperature.
2. PCR polymerases
Use a polymerase optimized for amplification of GC-rich sequences. To find an enzyme, visit our selection guide.