Fig. 4.

TrRad51 can process strand exchange with mismatched substrate. Fluorescence-based strand-exchange assay reveals a better mismatch tolerance of TrRad51 for mismatched substrate in strand exchange compared to ScRad51. (A) Schematic of the fluorescence-based strand-exchange assay. Successful strand-exchange events separate two fluorophores, resulting in increased Cy3 signal (see SI Appendix, Materials and Methods). (B) Schematics of homologous and mismatched substrates. (C) ScRad51 (I)- or ScDmc1 (II)-mediated strand exchange with homologous (blue) or mismatched (red) substrates was monitored by Cy3 emission signal at the indicated reaction time. Experimental repeats are presented as dotted curves and statistical analysis of all repeats is reported in E. (D) TrRad51-mediated strand exchange with all substrates was monitored by Cy3 emission signal at the indicated reaction time. (E) The mismatch tolerances of all recombinases with mismatched/homologous substrates were determined at 120 min. **P < 0.01; ****P < 0.0001. Data shown are average values ± SEM from three independent experiments. Statistics was performed by one-way ANOVA with Tukey’s post hoc test.