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. Author manuscript; available in PMC: 2014 Sep 8.
Published in final edited form as: Biochem Biophys Res Commun. 2012 Jun 17;424(1):40–44. doi: 10.1016/j.bbrc.2012.06.059

Figure 1. Effect of SpSsbB on SpRecA-promoted ATP-dependent three-strand exchange.

Figure 1

(A) Dependence on order of SpSsbB addition. The reaction solutions contained 5 µM circular φX ssDNA, 15 µM linear φX dsDNA, 5 mM ATP, 6 µM SpRecA, and no SpSsbB (minus SsbB), or 0.3 µM SpSsbB added 10 min after SpRecA (SsbB last), or 0.3 µM SpSsbB added 10 min before SpRecA (SsbB first). The reactions were carried out at 37 °C for the indicated times and then analyzed by agarose gel electrophoresis. In these reactions, the circular φX ssDNA (5 µM total nucleotide) was limiting relative to the linear φX dsDNA (15 µM total nucleotide = 7.5 µM base pairs) and therefore the maximum amount of linear dsDNA that could be converted to nicked circular dsDNA product was 67%. (B) Dependence on SpSsbB concentration. The reactions were carried out as described above (using the SsbB last order of addition) except that the concentration of SpSsbB was varied from 0 to 3.0 µM. The 60 min time points are shown. Labels: Slinear φX dsDNA substrate; I, partially exchanged intermediates; Pnicked circular φX dsDNA product; ssφX ssDNA substrate and product.