Figure 1.

Substrate design. A chimeric DNA/RNA oligonucleotide (5′-AAGTCrGACATCAG-3′) previously shown to be cleaved by MazF was labeled with 6-carboxyfluorescein (6-FAM) on the 5′-end and with Black Hole Quencher 1 (BHQ1) on the 3′-end. When the oligonucleotide is intact, BHQ1 quenches the fluorescence of 6-FAM; however, cleavage of the oligonucleotide at the RNA base by MazF releases 6-FAM from BHQ1, thereby increasing the fluorescence of 6-FAM.