A simple 'turn-on' fluorescent biosensor for sensitive detection of exonuclease III activity through photoinduced electron transfer and self-hybridization of a DNA probe

摘要

Exonuclease III (Exo III) plays crucial roles in maintaining the genome stability. Herein, a turn-on fluorescence strategy for detection of Exo III activity is developed. In this strategy, a poly(CG) single-stranded DNA (ssDNA) probe is labeled with fluorescein amidite (FAM) at the 3 end, and the ssDNA can self-hybridize to form double-stranded DNA (dsDNA). This dsDNA acts as the substrate for Exo III. Due to photoinduced electron transfer (PET) between FAM and guanine (G), the fluorescence of the 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 x 10(-4) U mL(-1) to 5 U mL(-1) with a detection limit of 3 x 10(-4) U mL(-1). 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.