They say megaprimer pcr, because one amplimer serves as a huge primer but with the double strain difference of a common primer.
I tried different approaches
A)30 cicles with only template and megaprimer.
B)Adding only the reverse primer in the first PCR in the annealing step after 5 cicles and continued 30 cicles
C)Adding both flanking primers in the first PCR in the annealing step after 5 cicles and continued 30 cicles
B and C is what they call Megaprimer Method.
when I use both flanking primers at 57 and 64 C° of Tanneal, I obtain a broad band near the expected size, as you can see in the picture
Then when I purified and try another pcr I do not have any kind of amplification.
Now I testing the changes in the annealing step, I use
Time I extend from 2 to 3 min
Tannealing from 44 to 52 °C
Tomorrow I´ll run the gel and tell you how it works..
Thank you so much for your help!