# Other than Taq? - for PCR after RT (Feb/15/2007 )

Hi,
do you know which one is the best to do PCR from cDNA of 3Kb?

The Taq doesn't seem to be efficient enough in this case!

thanks!

-biotech!-

If 3kb is the total size of the cDNA, then any high fidelity polymerase would do.

KOD hifi, Vent etc...

You don't need the really long template polymerase like phusion etc.

-perneseblue-

High Fidelity polymerase is very expensive though compared to Taq.

-timjim-

Ah but it will save you from falling to your knee screaming "Nooo!" when you find that your sequencing results show that nearly every single isolate you've sent has a basepair mutation somewhere.

Taq has an error rate somewhere around (1 x 10-4) to (2 x 10-5) errors per base pair.

Assuming (2 x 10-5) as the error rate per base pair
running 25 cycles
lenght 3000bp

The probability of an error in the fragment
P(E)
= fragment lenght * error rate per basepair
= 3000 * (2 x 10-5)
= 0.06

Probability that fragment has no error
=1-P(E)

Probability that after 25 cycles fragment still has no error
= [1-P(E)]^25
= [1-0.06]^25
= 0.21

Thus best case senario, only 1 in 5 fragment will be error free (21%). Worse case senario, the number is down to only 4%.

I would say it is definately better to spend the money buying a proper proof reading enzyme (avoid buying any "proof reading Taq"... the fidelity is better, but not by much.), it will save from trouble later.

-perneseblue-