PCR cycle for 80 bases - (May/04/2008 )
Can anyone suggest a PCR cycle to amplify 80 bases from a plasmid?
This depends more on the primers and enzyme than on the length of the amplified region. Without knowing anything else, I would try this:
95 2 min
35x (95 30s, 55 30s, 72 30s)
72 2 min
For extention time: You can find this information on Taq data sheet, depen on amplimer lenght.
Number of cycles: depen on concentration of your template.
HotStart: depen on what your Taq suport.
Annealing temperature . depen on your primers. I recommend to perform a gradient PCR.
If you use a normal PCR (not Realtime-PCR), the example of Phage434 it's fine. But you have to set up.!!! (maybe 30s of extention for 80 bp is too much, but you will find on Taq data sheet)