FAQ: I am having trouble amplifying a template that is longer than 5kb. How can I optimize my product yield using Phusion® High-Fidelity PCR Master Mix?

* Increase template amount. Sample concentration may be too low.
* Template DNA may be damaged. Use carefully purified template.
* Lengthen extension time.
* Increase the number of cycles.
* Lower annealing temperature.
* Titrate DMSO (2-8%) in the reaction.
* Denaturation temperature may be too low. Optimal denaturation temperature for most templates is 98°C or higher.
* Denaturation time may be too long or too short. Optimize the denaturation time.