2 replies to this topic

[anonymous]

I had used the dTTP to do the PCR before, but now I had changed to use the dUTP and the PCR product can not be generated. Does any one know how the PCR can work which the nucleotide used is dUTP.

[anonymous]

You have to be sure that the thermostable polymerase that you're using will incorporate dUTP into DNA, some enzymes like Pfx polymerase will not.  I would also suggest using double the concentrations of dUTP in your reactions since Taq tends to have a lower affinity for dUTP.  Good luck.

[stakie]

I have the same problem ... I can't get PCR products for AT rich sequences using dUTP instead of dTTP
I tried
* lowering annealing temperature
* Increasing dUTP concentration (twice as high as normal dNTPs)
* Trying different enzymes

Does anybody knows about an enzyme (or something else) that may solve this PCR problem?

Kind regards
Tim