I'm about to do a PCR cloning in which a 3.8Kb fragment is to be cloned using Clontech's In-Fusion kit. In the past I have used NEB Deep Vent and more recently Pfusion. Of those of you who do PCR cloning and need the efficient proof-reading function, which pol do you use?

Thanks

This may be a little outdated, but a few years ago a colleague of mine told me that she got best results using a mixture of two enzymes (1:1): AccuTaq (from Sigma) and Pfu (Fermentas). If this did not work, you can always spike some regular Taq (to get more amplicon). I assume that there are new proofreading enzymes out there that are even better, yet the idea I think still holds: if unsure of which enzyme is best and if it possible, try a mixture of enzymes. That way you may be able to harness the best of both worlds, assuming that each enzyme has an advantage over the other because of different reasons.

 I typically use Stratagene's PfuUltra II Fusion HS DNA polymerase. It's pretty fast, and it has great accuracy.

I don't need to have extra sensitivity because I'm using a plasmid as template. Does anyone else have a recommendation?

In that case, I would just use Pfu or Pfu Turbo from Stratagene (or any vendor, really).  Unless you're trying to amplify something very long, you should get plenty of amplification with 30-35 cycles of PCR using Pfu.  Use an extension time of 1 minute/kb.  I only mix Pfu with Taq if I have a long target from genomic DNA or cDNA.