Characterization of the DNA extension reaction. (A) Duplex-formation of published (black), suboptimal (green), and optimal (light green) oligonucleotide pairs. Numbers were obtained by quantification of electrophoretic mobility shift assays (EMSAs) shown in Supplemental Figure S4; SD (n = 3) are shown for each data point. (B) Relative DNA yields obtained in DNA polymerase extension reactions were quantified as described in the Materials and Methods. Errors, SD (n = 6), and P-values are shown. (C) PCR cycle-dependent amounts of extended DNA obtained by DNA polymerase reactions in the presence of 1 µM oligonucleotides and ∼10 nM Dream-Taq DNA polymerase (0.01 units) were quantified. The data were fitted to a single exponential curve. Errors, SD (n = 3) are shown. (D) The DNA template after 10 cycles of the DNA polymerase extension reaction was analyzed. After the DNA purification steps, DNA concentration was quantified by UV absorbance. Errors, SD (n = 3) are shown. (E) Time-dependent amounts of extended DNA obtained by DNA polymerase reactions in the presence of 1 µM oligonucleotides and ∼10 nM Dream-Taq DNA polymerase (0.01 units) were quantified. Errors, SD (n = 3) are shown. (F) DNA yields after 25 cycles of the extension reaction in the presence of ∼10 nM and ∼250 nM Dream-Taq DNA polymerase were analyzed after DNA purification steps. DNA concentration was quantified by UV absorbance. Errors, SD (n = 3) are shown.
