Figure 2.

A schematic diagram of molecular beacons, molecular break lights, and the specific break lights used in this study. The solid lines represent covalent bonds, dashed lines represent hydrogen bonding, letters represent arbitrary bases, the gray shaded ball represents the fluorophore (FAM), the black ball represents the corresponding quencher (DABCYL), and the dashed wedges represent fluorescence. (a) Principle of operation of molecular beacons. Target hybridization leads to a separation of the fluorophore–quencher pair and a corresponding fluorescent signal. (b) Principle of operation of molecular break lights. Cleavage of the stem by an enzymatic or nonenzymatic nuclease activity results in the separation of the fluorophore–quencher pair and a corresponding fluorescent signal. (c) Molecular break lights used in this study. The stem of break light A contains a preferred calicheamicin recognition site (in bold), and the stem of break light B carries the BamHI recognition site (in bold). The predicted cleavage sites are illustrated by arrows.