High fidelity DNA synthesis by the Thermus aquaticus DNA polymerase.

Abstract:

We demonstrate that despite lacking a 3'----5' proofreading exonuclease, the Thermus aquaticus (Taq) DNA polymerase can catalyze highly accurate DNA synthesis in vitro. Under defined reaction conditions, the error rate per nucleotide polymerized at 70 degrees C can be as low as 10(-5) for base substitution errors and 10(-6) for frameshift errors. The frequency of mutations produced during a single round of DNA synthesis of the lac Z alpha gene by Taq polymerase responds to changes in dNTP concentration, pH, and the concentration of MgCl2 relative to the total concentration of deoxynucleotide triphosphates present in the reaction. Both base substitution and frameshift error rates of less than 1/100,000 were observed at pH 5-6 (70 degrees C) or when MgCl2 and deoxynucleotide triphosphates were present at equimolar concentrations. These high fidelity reaction conditions for DNA synthesis by the Taq polymerase may be useful for specialized uses of DNA amplified by the polymerase chain reaction.

Polymerases:

Topics:

Fidelity, Exonuclease Activity, Source / Purification

One line summary:

This paper describes high fidelity reaction conditions for DNA synthesis by the Taq polymerase.

Status:

new topics/pols set partial results complete validated

Results:

Polymerase Reference Property Result Context
Taq pol I Eckert KA1990 3-5' Exonuclease (proofreading) No
Taq pol I Eckert KA1990 Cloned or native Cloned in E. coli
Taq pol I Eckert KA1990 Frameshift Error Rate 5.26E-05 errors/bp Technique: Reversion (Low fidelity conditions)
Taq pol I Eckert KA1990 Frameshift Error Rate 4.76E-06 errors/bp Technique: Reversion (High fidelity conditions)
Taq pol I Eckert KA1990 Nucleotide Substitution Rate 0.0003125 errors/bp Technique: M13mp2 opal codon reversion assay (Low fidelity conditions)
Taq pol I Eckert KA1990 Nucleotide Substitution Rate 3.33E-06 errors/bp Technique: M13mp2 opal codon reversion assay (High fidelity conditions)
Taq pol I Eckert KA1990 Full length or truncated Full length

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