Figure 1.

P. abyssi RadA recombinase activity catalyzed displacement-loop (D-loop) formation. (A) Schematic representation for the D-loop formation assay. Labeled linear ssDNA (93 nt) was incubated first with RadA to form nucleoprotein filaments before adding the purified supercoiled plasmid pUC19 for further homology search. (B) D-loop formation assay with increased quantity of RadA. An amount of 25 nM of labeled ssDNA (93 nt) was incubated with RadA for 10 min at 65 °C. Then, 25 nM of purified supercoiled pUC19 was added and incubated for another 10 min. DNA products were separated on a 1.2% native agarose gel and visualized by fluorescence. (C) Histogram representation of the D-loop formation assays for a range of RadA as observed in (B). RadA-dependent D-loop (%), densitometry measurement of formed D-loop as a percentage of total lane densitometry after data normalization and the D-loop background from lane 3 was subtracted. Experiments were performed in triplicate.