RNase H amplified RNA probe and graphene oxide system for highly sensitive detection of (CAG)n DNA repeat sequences

Abstract

Huntington’s disease is a chronic progressive neurodegeneration which is caused by CAG repeat sequences expanding in the huntingtin gene. There is currently no disease-modifying treatment for the disease, and its progression can only be slowed down before the onset of symptoms. A novel fluorescent platform which contains an RNA probe and graphene oxide for detection of the biomarker of Huntington’s disease, CAG repeat sequences, was constructed in this investigation. In addition, RNase H was employed in the fluorescent system to enhance the sensitivity of the detection capability. The fluorescent signal was increased through the cyclic amplified reaction, which results from RNase H, specifically digestion of the RNA strand in the complement of the RNA–DNA duplex. The designed measurement method can detect CAG repeat sequences with a detection limit of 108 pM (R2 = 0.968) under which we optimized assay conditions. Furthermore, the detection limit is approximately 18 times lower than the traditional DNA and graphene oxide detection method without assistance of RNase H. Additionally, the probing platform also shows stronger ability to discriminate between the fluorescence of the target sequence and that of other non-target sequences. The results of our studies demonstrate that the RNase H amplified RNA probe and graphene oxide system exhibited excellent sensitivity and selectivity to the target of CAG repeats sequences.