Moreover, to investigate whether TcaR preferentially binds to ssDNA or dsDNA, the ability of ssDNA to compete with the TcaR-dsDNA complex was evaluated. For the competition assay, the IcaR DNA1 probe was preincubated with TcaR (dimer) protein to allow formation of the dsDNA-TcaR complex prior to mixing with increasing amounts of single-stranded GC33 DNA. It has been known that ssDNA products have lower migration velocity compared to its dsDNA counterparts in polyacrylamide electrophoresis [24, 25] . As shown in Figure 1E , ssDNA, as a competitor, interfered the binding of TcaR to the dsDNA, suggesting a binding preference for ssDNA. To further confirm this result, IcaR DNA1 and GC33 ssDNA oligomers were mixed, and their interaction strengths with TcaR were compared using EMSA ( Figure 1F ). Findings indicated that increasing the concentration of TcaR produces a ssDNA band shift greater than that for dsDNA, confirming a stronger interaction between TcaR and ssDNA. Moreover, to investigate possible pH effect of ssDNA binding activity of TcaR, a series of buffers with increasing pH were tested for their potential interfere in TcaR-ssDNA binding. As shown in Figure 1G , the EMSA results showed that TcaR had a strongest affinity for GC33 at pH 8.0 and the affinity was reduced by decreasing pH. Consequently, the result indicates that the ssDNA binding activity of TcaR is pH-dependent.