Troubles with Pfu polymerase

[Koen De Smet]

hatzfeld at msdos.ensam.inra.fr (Yves hatzfeld) wrote:
>Hi Netters!
>
>I have troubles with amplifying a 1.5 kb fragment with pfu polymerase
>(stratagene). 
[...deleted some bits....]
>When I do the same experiments using Taq instead, I haven't any problem
>and I obtain the expected products ...
>
>So why the Hell it does not work? I need to use pfu because I have to
>
>Yves.

I recently emailed Stratagene with a related question on pfu polymerase. This is the answer I got from them:

     Dear Dr.  De Smet,
     
     pfu polymerase has amplified up to 12 kb, so 3 kb is not a problem.  I 
     should, however, mention that pfu sometimes requires more optimization 
     that taq.  Start by using a 40C annealing temp. and 2-3 minutes per kb 
     extention times for at least 30 cycles.
     Best regards,
     
     Will Scovill

Maybe this helps? 
Koen De Smet
<A HREF="hhtp://www.sm.ic.ac.uk/medmicro/home.htm">Imperial College Medical School at St  Mary's, London</A>