Figure 4: Cofactor requirement and gel shift assay. (a) The purified protein (2 μg) was incubated with supercoiled pBluescript II KS (1 μg) in buffer R with (+) and without (−) 10 mM MgCl2 for 60 min at 37°C before being analyzed on an agarose gel. (b) The purified 500 bp fragment (1 μg) was incubated with increasing amount of protein in buffer R (total volume 50 μL) for 15 min at 37°C and was analyzed on 1.2% agarose gel. Lanes 1 and 5 are size standard of a purified 500 bp DNA fragment. The 500 bp fragment (1 μg) was incubated with the purified enzyme at 5 μg (lane 2), 7 μg (lane 3), and 8.8 μg (lane 4), respectively.