Eckert K A, Kunkel T A
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709.
Nucleic Acids Res. 1990 Jul 11;18(13):3739-44. doi: 10.1093/nar/18.13.3739.
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.
我们证明，尽管嗜热水生栖热菌（Taq）DNA聚合酶缺乏3'→5'校对核酸外切酶，但它在体外仍能催化高度精确的DNA合成。在特定反应条件下，70℃时每聚合一个核苷酸的碱基替代错误率可低至10^(-5)，移码错误率可低至10^(-6)。Taq聚合酶在对lac Zα基因进行单轮DNA合成过程中产生的突变频率会随dNTP浓度、pH值以及MgCl2浓度相对于反应中脱氧核苷酸三磷酸总浓度的变化而改变。在pH 5 - 6（70℃）时，或者当MgCl2和脱氧核苷酸三磷酸以等摩尔浓度存在时，碱基替代和移码错误率均低于1/100,000。Taq聚合酶进行DNA合成的这些高保真反应条件可能对聚合酶链反应扩增的DNA的特殊用途有用。
