Effect of heterologous RPA on the coupled DNA pol α–primase activity on ssM13 DNA. (A) The coupled DNA pol α–primase reactions containing 40 ng of unprimed M13 ssDNA template and 0.1 U of human pol α–primase complex were incubated at 37°C for 30 min in the presence of 0.4 µg RPA (except lane 1) and the increasing amount of T-ag (0 µg, lanes 1, 2, 5, 8, 11, 14, 17, 20 and 23; 0.4 µg, lanes 3, 6, 9, 12, 15, 18, 21 and 24; 0.8 µg, lanes 4, 7, 10, 13, 16, 19, 22 and 25). The heterologous RPAs used are indicated by the three-letter symbol. After incubation, reaction products were isolated as described in Materials and Methods and analyzed by 20% PAGE containing 7 M urea, 1 mM EDTA and 89 mM Tris–borate (pH 8.0). (B) For quantitation, regions containing reaction products were excised and analyzed by liquid scintillation counting.