BioMed Research International / 2014 / Article / Fig 4

Research Article

C-Terminal Domain Swapping of SSB Changes the Size of the ssDNA Binding Site

Figure 4

Binding of KpSSB to dT20–60. KpSSB (0, 19, 37, 77, 155, 310, 630, 1250, 2500, and 5000 nM) was incubated for 30 min at 25°C with 1.7 nM of (a) dT20, (b) dT25, (c) dT35, (d) dT45, (e) dT50, (f) dT55, or (g) dT60 in a total volume of 10 μL in 20 mM Tris-HCl pH 8.0 and 100 mM NaCl. Aliquots (5 μL) were removed from each reaction solution and added to 2 μL of gel-loading solution (0.25% bromophenol blue and 40% sucrose). The resulting samples were resolved on a native 8% polyacrylamide gel at 4°C in TBE buffer (89 mM Tris borate and 1 mM EDTA) for 1 h at 100 V and visualized by autoradiography. Complexed and free DNA bands were scanned and quantified.
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