The fidelity of Taq polymerase catalyzing PCR is improved by an N-terminal deletion.
KlenTaq DNA polymerase is an N-terminally truncated Thermus aquaticus (Taq) DNA polymerase I. As expressed from a gene construct in Escherichia coli, translation initiates at Met236, bypassing the 5'----3' exonuclease domain of the DNA polymerase-encoding gene. A sensitive forward mutation assay was used to measure the relative number of mutations introduced into the entire lacZ gene by the polymerase chain reaction (PCR) under various conditions which allow the amplification of such a large DNA span. Two selectable markers, one at each end of the test lacZ fragment, were employed to avoid the plating and scoring of PCR artefacts such as primer initiation in the midst of the lacZ gene, and cloning artefacts such as empty vector plasmid. The measured relative mutation rate was twofold lower for KlenTaq as compared to the full-length Taq DNA polymerase.
Barnes
W M
WM
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO 63110.
D001483:Base Sequence; D003001:Cloning, Molecular; D004274:DNA, Recombinant / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D007763:Lac Operon; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D019914:Taq Polymerase; D013824:Thermus / Q000201:enzymology*; D001616:beta-Galactosidase / Q000235:genetics
D004274:DNA, Recombinant; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase; D001616:beta-Galactosidase