Simple “turn-on” fluorescent biosensor for sensitive detection of exonuclease III activity through photoinduced electron transfer and self-hybridization of one DNA probe

Qiang Liu, Jinyu Lian, Mei Liu, Yan Jin, Baoxin Li

Index: 10.1039/C8AY00631H

Full Text: HTML

Abstract

Exonuclease III (Exo III) plays crucial roles in maintaining the genome stability. Herein, a turn-on fluorescent strategy for detection of Exo III activity is developed. In this strategy, one poly(CG) single-stranded DNA (ssDNA) probe is labeled with fluorescein amidite (FAM) at 3′ end, and the ssDNA can self-hybridize to form the double-stranded DNA (dsDNA). This dsDNA acts as the substrate of Exo III. Due to photoinduced electron transfer (PET) between FAM and guanine (G), the fluorescence of FAM-labeled dsDNA probe is completely quenched. Upon the addition of Exo III, the dsDNA will be digested, and FAM emits very strong fluorescence. Thus, Exo III activity can be facilely measured with a simple fluorescence reader. This method has a wide detection range from 5×104 U/mL to 5 U/mL with a detection of 3×10-4 U/mL. The results demonstrated herein may provide a new pattern for Exo III activity detection with high accuracy and good specificity as well as satisfactory applicability in real biosamples, which holds great potential for drug screening and basic research related to exonucleases.