Two annealing temps? - (Sep/17/2009 )
Hi all, I'm using someone else's PCR protocol - there are two annealing temps, first 48deg for 20 sec then 55deg for 20 sec. Any idea why there are two temps? Also since I'm seeing non-specific amplification (and I'd usually try upping the annealing temp to get rid of it) any suggestions on which annealing temp to up?
microgirl on Sep 17 2009, 09:48 AM said:
It sounds to me like a touchdown PCR...Is that 48 degree for 20sec, 10 cycles, then 55deg for 25 cycles..something like that.?
xiaoposhi on Sep 17 2009, 12:20 PM said:
In touchdown temperature is decreased in every cycle, not increased.
It sounds to me like a touchdown PCR...Is that 48 degree for 20sec, 10 cycles, then 55deg for 25 cycles..something like that.?