Do I Need to Work on Ice for Hot Start PCR?

[sotellme]

Do i need to work on ice when i do Hot start PCR?


Hope for inputs.

Thanks.

 

[Moonbear]

As a general rule, I work with everything on ice for PCR unless the protocol specifies otherwise.

 

[Monique]

Yes, I always keep everything on ice too (best to be on the safe side )

 

[iansmith]

You do not have to work on ice for HS-PCR. The idea of working on ice when you are not doing HS-PCR is to reduce the activity of the polymerase and to reduce unspecific annealing of the primers. This is usually sufficient to elimination most of the unspecific binding problems.

For HS-PCR, the unspecific binding are eliminated because any unspecific annealed primers has been removed during first denaturation step. So when you add Taq after the denaturation step, unspecific annealed primers should not be present.