DNA Tetrahedron Based Biosensor for Argonaute2 Assay in Single Cells and Human Immunodeficiency Virus Type-1 Related Ribonuclease H Detection in Vitro.

Abstract

A well-designed DNA tetrahedron based biosensor (DTB) has been employed for imaging and detection of argonaute2 (Ago2), a key RNA interference (RNAi) protein. This DTB mainly contains two segments: DNA tetrahedron as a frame and a photoinduced electron-transfer (PET) pair as a fluorescence transducer. The DNA tetrahedral nanostructure is assembled with four nucleic acid strands. On one edge, one DNA strand forms a hairpin structure (RNA sequence) which is implanted in the two complementary double-strand DNAs, and the PET pairs, DNA/silver nanocluster (AgNC) and G-quadruplex/hemin complex, are labeled at the two termini of another DNA strand, respectively. The DNA tetrahedron structure forms a switchable scaffold that can present the functional DNA motifs in two different modes, according to the presence/absence of the target protein. The cleavage reaction by Ago2/miR-21 complex opens the hairpin structure, leading the PET pairs to be separated to each other in the spatial state. Thus, the DNA/AgNC fluorescence can be measured. By using this DTB, we obtained the 4.54 nM Ago2 detection limit and successful assay of Ago2/miR-21 complex in living cells. Also, the DTB was successfully employed for the further assay of RNase H, which plays an important role in the human immunodeficiency virus type-1 (HIV-1) reverse transcriptase pathway, with a limit of detection (LOD) of 3.41 U mL-1. Our DTB can be used as a device for the imaging protein concentration in single cells and has a potential value for disease diagnosis and treatment.