Help! pfu DNA polymerase for PCR

Yi, Xiaoming Z3C20 at ttacs3.ttu.edu
Wed May 10 17:33:44 EST 1995


Pei Wu,

I had the same problem before(Taq works but pfu does not). One suggestion I
have is use another buffer: make 100mM Tris base(use sulfuric acid to adjust pH
to 8.75), 100mM (NH4)2SO4, 70mM MgSO4. This is 10X buffer, try this if your
primers are working with Taq.

Xiaoming Yi
Texas Tech University


In <johnaris-1005951308500001 at 128.227.67.104> johnaris at anatomy.med.ufl.edu writes:

> Dear colleagues,
> 
> I have been using the pfu DNA polymerase to try to amplify a 3-kb fragment, I
> tried different conditions such as annealing temperatures at 45, 50, 55, 60
> degrees, extension time 6, 7 and 8 minutes at 72 degrees. I am using the yeast
> genomic DNA as template, I tried 10 and 100ng per 50 ul reaction and I also 
> tried 2.5 and 5 units per 50 ul reaction, each time I used taq polymerase as
> a control. I can obtain a 3-kb product from taq but nothing from pfu. If
> anybody knows how pfu works, would you please tell me about it? Thanks.
> 
> Pei Wu
> Anatomy and Cell Biology
> University of Florida
> (904) 392-4563



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