Radiolabeled annealed primer/template (250 nM). Position 1 in each template (lanes 2−5) provided the corresponding Watson−Crick complementary deoxynucleoside to test elongation of the DNA primer by one residue. Each reaction included 20 mM Tris-HCl, 10 mM KCl, 10 mM (NH4)2SO4, 20 mM MgSO4, 0.1%
Triton X-100, 0.2 μg/μL BSA, 100 μM DTT, 0.2 units (37 nM) of
Therminator Polymerase, 1.25 mM Mn2+, and 0.5 μM tNTP and
were incubated at 55 °C for 15 min. Lane 1: Radiolabeled primer.
Lane 2: tGTP (no Mn2+). Lane 3: tDTP. Lane 4: tTTP. Lane 5:
tCTP. Lane 6: tGTP, tDTP, tTTP, and tCTP with an incubation
time of 120 min.