Posted 13 February 2012 - 11:10 PM
It is described in the attachment...
Posted 14 February 2012 - 03:40 AM
Is this your first amplification or a "recipe" passed down to you?
Reason I asked is that the final concentration of Mg2+ (3mM) is quite high for me. For Taq, I usually add it into the individual tubes before starting the cycles. I add the primers into the master mix though.
I have no complains about the cycles and temperature but depending on the outcome, the annealing time [and temperature] might change.
Unless this is a repeat of what was done before, you might need some tweaking after this.
Edited by Julio-Claudian, 14 February 2012 - 03:58 AM.
Posted 14 February 2012 - 08:41 PM
Posted 14 February 2012 - 09:15 PM
Since it's a passed down secret recipe, I'm sure it's been optimised (?). So I think there must be a reason for such values. In any case, perform the amplification, see what you get, and work from there.
I remember starting my Mg2+ at 1.75 mM and [together with other factors] went up until 2.5 mM. But like I said, it depends.