Figure 3.

Dmc1 reactions lacking RPA are more efficient when Hop2-Mnd1 and Rdh54/Tid1 are added late to reactions. (A) The order of addition of reaction components is as indicated. The concentrations of reactants are as detailed in Figure 1 and in Materials and Methods. The first arrow indicates incubation was at 37°C for 10 min followed by the second arrow, which indicates an additional incubation at 37°C for 15 min. Hop2-Mnd1 and dsDNA in reaction 1 were incubated at room temperature for 3 min prior to adding to Dmc1-ssDNA filaments. The gel images from D-loop assays are shown and results graphed (n = 3, ± s.e.m.). Data shown here was published previously (16) and are presented for comparison to B with permission [NB: permission pending]. (B) The same experimental procedure as in A except Rdh54/Tid1 was used in place of Hop2-Mnd1. (C) Deletion and add-back reactions in the pre-mixed regimen of Hop2-Mnd1 or Rdh54/Tid1 in the absence of RPA. The order of addition of components were as indicated. Abbreviations for proteins and protein and DNA concentration are as in Figure 1.