Effect of salts, solvents and buffer on miRNA detection using DNA silver nanocluster (DNA/AgNCs) probes

Abstract

MicroRNAs (miRNAs) are small regulatory RNAs (size ~21 nt to ~25 nt) which regulate a variety of important cellular events in plants, animals and single cell eukaryotes. Especially because of their use in diagnostics of human diseases, efforts have been directed towards the invention of a rapid, simple and sequence selective detection method for miRNAs. Recently, we reported an innovative method for the determination of miRNA levels using the red fluorescent properties of DNA/silver nanoclusters (DNA/AgNCs). Our method is based on monitoring the emission drop of a DNA/AgNCs probe in the presence of its specific target miRNA. Accordingly, the accuracy and efficiency of the method relies on the sensitivity of hybridization between the probe and target. To gain specific and robust hybridization between probe and target, we investigated a range of diverse salts, organic solvents, and buffer to optimize target sensing conditions. Under the newly adjusted conditions, the target sensitivity and the formation of emissive DNA/AgNCs probes were significantly improved. Also, fortification of the Tris-acetate buffer with inorganic salts or organic solvents improved the sensitivity of the DNA/AgNC probes. On the basis of these optimizations, the versatility of the DNA/AgNCs-based miRNA detection method can be expanded.

DNA/RNA chimera templates improve the emission intensity and target the accessibility of silver nanocluster-based sensors for human microRNA detection

“…To generalize the DNA/AgNC-based method, we further investigated the mechanism underlying the emission of strong fluorescence and optimized it under buffer conditions. 38,39 Furthermore, we showed that multiplex detection of miRNAs in solution is highly feasible using different DNA/AgNC sensors with different emission spectra. 40 In fact, we demonstrated that the method is practical enough for screening a novel mutant plant, whose miRNA biogenesis pathway is notably compromised or disrupted.…”

The role of spacer sequence in modulating turn-on fluorescence of DNA-templated silver nanoclusters

“…The use of DNA template has the advantage of achieving specific target recognition through the direct incorporation of complementary DNA or aptamer sequences without the need for post-synthesis functionalization ( 37–40 ). There has been increased understanding on how the structure of AgNC correlates to its fluorescence properties ( 41–45 ); though for AgNC–DNA to truly take off as a toolbox for biomedical applications, the next technical hurdle is to generate DNA sequences for templating AgNC–DNA with good stability particularly in a chloride-rich environment ( 46 ). Some strategies were previously proposed to enhance the stability of AgNC–DNA, e.g.…”

Dengue serotyping with a label-free DNA sensor

“…In addition to this, certain salts such as chloride ions have also been reported to interfere with AgNC formation when present at high concentrations. 59 Chloride ions have been shown to obstruct the formation of AgNCs via AgCl precipitation or the sequestering of silver ions. 59 Hence, chloride ions were replaced with nitrate ions in the preparation of Buffer A and Buffer B for the assay to avoid such issues.…”