error rate of polymeases
simon.andrews at bbsrc.ac.uk
Tue Apr 10 04:05:13 EST 2001
Wolfgang Schechinger wrote:
> Hi all,
> To my knowledge, the error rate of Taq is said to be 1/1000 ,
> that of proofreading Pols (Pwo, Pfu) ist said to be 1/5000.
> Are these figures close to reality and what do they actually
> mean? When I copy a gene with Taq,
> there will be (statistically) 1 error in every 1000 bases?
> Meaning that after a 30 cycle (theoretical) PCR starting from a
> single copy, there should be 30 errors within 1000 bp (after
> 30 duplications)?
There are many quoted values for the fidelity of polymerases. The
values you quote above are way to high to be mistakes per base
transcribed. Remember PCR is an exponential process so that mistakes
made in round 1 get replicated through subsequent rounds. A polymerase
writing 1 in 1000 bases wrong would make some seriously mangled products
after 30 rounds of amplification!! I suspect those values are average
mistakes per product following amplification. I seem to remember that
mistakes per base generally runs at about 1 in 10^6 for Taq.
The overall fidelity depends on many factors including amount and
composition of template, concentration of dNTPs, Mg concentration,
number of cycles run etc. etc.
There are numerous papers on this subject - one reference I have to hand
Keohavong, P et al. (1989) Fidelity of DNA polymerases in DNA
amplification. PNAS-USA 86:9253-9257
Hope this helps
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