A dual-signal amplification method for the DNA detection based on exonuclease III

A dual-signal amplification method based on two molecular beacons was designed for human hemochromatosis (HFE) gene detection. The two probes, P-1 and P-2, could resist the exonuclease III (Exo III) digestion due to the 3 '-termini protrusion, and could coexist stably with Exo III. In the presence of HFE targets, P-1 hybridized with a HFE target to form a duplex DNA with a recessed 3 '-hydroxyl termini and then partially digested by Exo III, releasing the HFE target and a residual sequence (X). This X sequence could trigger the digestion of P-2 probes with 6-carboxy-fluoresceins and Black Hole Quenchers and then result in the increase of fluorescence intensity. The X sequences were more stable than HFE targets and could cyclically trigger the P-2 digestion for a long time even though the HFE targets were digested by Exo III. This method improved the sensitivity and reached 4 orders of magnitude in detection limit, and showed excellent selectivity to discriminate single base mismatched targets well. (C) 2014 Elsevier B.V. All rights reserved.