Fig. 3.
Truncated elongation on Nf wild-type and mutated single-stranded oligonucleotide templates. The TP-linked DNA molecule resultant of replicating ori(29)Nf in the presence of ddCTP is shown, assuming that the first and second nucleotides are recovered during elongation. Underlined T in the template molecule marks the initiation site. Reactions were carried out in the presence of 2.3 μM the indicated oligonucleotide, 60 nM Nf DNA polymerase, 120 nM Nf TP, 0.1 μM the corresponding labeled dNTP, 10 μM each dATP, dGTP and dTTP, and 100 μM ddCTP. Reactions were started by adding 1 mM MnCl2 and, after 30 min at 30 °C, were stopped and processed as indicated in Materials and Methods. In addition to the initiation complex (TP-dAMP or TP-dTMP), the position corresponding to different end-products (TP-(dNMP)n) of each truncated elongation assay is indicated.