FIGURE 7.

FEN1 processing of substrates containing flaps of 60 nt under single-turnover conditions. A, representative time courses of FEN1 acting upon 60M (filled triangles) and 60R (open triangles). B, representative autoradiograms depicting cleavage of 60M (top panel) and 60R (bottom panel). The 0 reaction time is a negative control (see “Experimental Procedures”). The zoomed in box to the right of 60R depicts the ladder-type pattern of cleavage. Experimental conditions were 10 nm DNA substrate, 50 nm FEN1, 30 mm HEPES-KOH, 40 mm KCl, 8 mm MgCl₂, 5% (v/v) glycerol, 0.1 mg/ml BSA, pH 7.4. Reactions were performed at 37 °C using an RQF instrument and quenched with EDTA.