Hello,

I am wanting to do overlap PCR on two fragments I have. I want to use the strategy where the overlap region acts as the primers for extending the fragment.

For the overlap PCR, what ratio and how much DNA should I add to the overlap reaction? Second, using phusion polymerase, what is a common protocol I can use for annealing the overlap area, and extending? I know this can differ depending on how much extension is required. By I rough idea on how the protocol would look like would be great.

Thanks