In-solution multiplex miRNA detection using DNA-templated silver nanocluster probes

Abstract

MicroRNAs (miRNAs) are small regulatory RNAs (size ∼21nt to ∼25nt) that can be used as biomarkers of disease diagnosis, and efforts have been directed towards the invention of a rapid, simple and sequence-selective detection method for miRNAs. We recently developed a DNA/silver nanoclusters (AgNCs)-based turn-off fluorescence method in the presence of target miRNA. To further advance our method toward multiplex miRNA detection in solution, the design of various fluorescent DNA/AgNCs probes was essential. Therefore, tethering of DNA-12nt scaffolds with 9 different AgNCs emitters to target-sensing DNA sequences was investigated. Interestingly, for the creation of spectrally different DNA/AgNCs probes, not only were the emitters encapsulated in 9 different DNA-12nt scaffolds necessary but the tethered target-sensing DNA sequences are also crucial to tune the fluorescence across the visible to infra-red region. In this study, we obtained three spectrally distinctive emitters of each DNA/AgNCs probes such as green, red, and near-infrared (NIR) fluorescence. Using these DNA/AgNCs probes, we here show a proof of concept for a rapid, one-step, in-solution multiplex miRNA detection method.

COP1 E3 ligase protects HYL1 to retain microRNA biogenesis

“…For the miRNA detection assay, we mixed a fixed amount of total RNAs (5 mg) from cop1-6, hyl1-2 and wild type with the probes at the given concentrations of Tris-acetate buffer and incubated for 15 min at 25°C. Then, AgNO 3 (250 mM) and NaBH 4 (250 mM) were added to the RNA/DNA mixtures to a final volume (50 ml) (14)(15)(16)(17). All the DNA/AgNCs reactions were incubated for 1 h at 25°C and diluted with 450 ml of distilled water before measurement on a fluorimeter (Horiba Jobin Yvon, Fluoromax-4) in a 10 mm disposable cuvette.…”

“…In addition to the core processing proteins, at least 10 components have been reported that play direct or indirect roles in the intricate regulation of miRNA biogenesis [7][8][9][10][11][12][13] . To further isolate hidden components in miRNA biogenesis, we have developed a rapid and simple miRNA detection method using DNAtemplated silver nanoclusters (DNA/AgNCs) probes [14][15][16][17] . By applying this method, we isolated COP1 E3 ubiquitin ligase that negatively regulates photomorphogenesis triggered by far-red and visible light.…”

miRNA Optical Detection

“…Numerous studies have applied DNA as a scaffold for nanocluster formation, focusing on the photoluminescence aspects of the DNA-templated silver nanoclusters (DNA/AgNCs) for biosensing application [22][23][24]. Bjerrum and Yang [25] have developed a strategy based on various fluorescent DNA/AgNCs probes for multiplex miRNA detection in solution. For the creation of spectrally different DNA/AgNCs probes, the emitters are encapsulated in nine different DNA12nt scaffolds, which is necessary, and with the tethered target-sensing DNA sequences (also crucial) to tune the fluorescence across the visible to infrared region ( Fig.…”

Nanomaterials-Based Fluorimetric Methods for MicroRNAs Detection

“…When target miRNAs are present, the emission of the DNA-nanosilver clusters (DNA/AgNCs) probe is significantly lower versus the case when no target miRNAs or other miRNAs are present. Additionally, to further advance the method toward multiplex miRNAs detection in solution, they also presented the design of three DNA/AgNCs probes showing green, red, and near-infrared (NIR) fluorescence [20]. Besides, Ye and co-workers reported the quantification of miRNAs expression levels in cell lysates by target assisted isothermal exponential amplification (TAIEA) coupled with fluorescent DNA-scaffolded AgNCs [16,21].…”