Figure 3.
The strand displacement activity of Pol δDV is inhibited by addition ‘in trans’ of ssDNA. (a) Scheme depicting the experimental strategy. After pre-forming a Pol δDV–DNA complex the reactions are started either by (1) adding dNTP, (2) adding dNTP and ssDNA or (3) first adding ssDNA to the pre-formed complex followed by addition of dNTP. (b) Effect of different concentrations of dT25 on the primer extension and strand displacement activities of Pol δDV in Buffer TM (20 mM NaCl, 30°C) using either a labeled, primed DNA template or a labeled T7 gap substrate without a 5′-flap in the strand to be displaced. (c) ssDNA length dependence of the ‘in trans’ inhibition using a 10-fold excess of dT18, dT14 and dT10. For dT10 experiments using a 20-fold excess are shown as well. The boxed numbers in (b) and (c) refer to the way the experiments were performed according to (a).