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. Author manuscript; available in PMC: 2013 Dec 21.
Published in final edited form as: Biochemistry. 2012 Dec 11;51(51):10147–10158. doi: 10.1021/bi301202e

Figure 6. Gap filling by PolD2 extends into the downstream duplex segment.

Figure 6

(A) Reaction mixtures (20 µl) containing 50 mM Tris-HCl (pH 8.0), 1 pmol of 32P-labeled DNA substrate (gapped duplexes or simple primer-template as illustrated above the lanes), 100 µM each of dATP, dGTP, dCTP and dTTP (dNTPs), 1.25 mM CoCl2, and MsmPolD2 as indicated above the lanes were incubated for 10 min at 37°C. (B) Reaction mixtures (140 µl) containing 50 mM Tris-HCl (pH 8.0), 100 µM each of dATP, dGTP, dCTP and dTTP (dNTPs), 1.25 mM CoCl2,, 7 pmol of 32P-labeled substrate (simple primer-template or gapped duplex as illustrated below the lanes), and 1.2 µg of MsmPolD2 were incubated at 37°C. Aliquots (20 µl) were withdrawn at the times specified and quenched immediately with EDTA/formamide. The products were analyzed by PAGE and visualized by autoradiography.