Electrophoretic mobility shift assays.

The DNA fragment of the arsR2 regulatory region was amplified using the primers arsK-EMSA-F and arsK-EMSA-R (Table 2). The forward primer was labeled with the fluorophore 6-carboxyfluorescein (FAM). All reaction mixtures with or without metalloids were incubated at 28°C for 30 min in binding buffer (20 mM Tris-HCl [pH 7.0], 50 mM NaCl, 1 mM dithiothreitol [DTT], 10 mM MgCl₂, 100 µg/ml bovine serum albumin [BSA]). The binding solution was then loaded onto a 6% native PAGE gel. After 3 h of running at 80 V in 1× Tris-glycine-EDTA (TGE) buffer (120 mM Tris, 950 mM glycine, 5 mM EDTA), the gels were exposed in a phosphor imaging system (Fujifilm FLA-5100) (31).