Self-Priming DNA Polymerization-Propelled Stochastic Walkers on Magnetic Microbeads for Amplified Detection of miRNA

Abstract

Sensitive detection of miRNA targets in complex biological samples possesses great value in biopsy analysis and disease diagnosis but is still challenging because of low abundance and nonspecific interferences. In this work, self-primer DNA polymerization-propelled stochastic walkers (SWs) were proposed to detect miRNA-24 by combining magnetic microbeads (MMBs) and flow cytometry. The MMBs not only provide a three-dimensional interface for DNA walkers but also facilitate the enrichment and isolation of RNA targets from complex biological samples such as serum. The SWs can be initiated to walk through the entire surface of MMBs and transduce RNA walking into amplified fluorescence signals, with the detection limit of miRNA-24 at 0.95 pM. Moreover, this strategy integrating with flow cytometry was demonstrated to have good specificity with other homologous miRNAs. This platform offers promising applications in RNA biosensing and biomedical diagnostics.

Corn-Based Fluorescent Light-Up Biosensors with Improved Signal-to-Background Ratio for Label-Free Detection of Long Noncoding RNAs

“…In summary, we have constructed a new Corn-based light-up biosensor for label-free detection of lncRNA by integrating DSN-assisted target recycling amplification with transcription-driven synthesis of fluorogenic RNA aptamer. This Corn-based biosensor has distinct merits: (1) the introduction of magnetic bead-capture probe conjugates facilitates efficient enrichment and rapid separation of target lncRNA from complex biological matrices, greatly improving assay speed and reducing background signal; (2) target lncRNA signal is rapidly amplified into the T7 promoter signal through a DSN-assisted cyclic cleavage reaction without the involvement of converting lncRNA into its cDNA, efficiently eliminating the contamination of genomic DNAs; (3) unlike DNA polymerase-catalyzed amplification reaction with inevitable shortcoming of nonspecific background amplification induced by the primer/template-independent polymerization, T7 RNA polymerase-based transcription amplification is extremely specific T7 promoter-dependent to generate RNA sequences, greatly reducing non-specific amplification and improving detection accuracy; (4) the introduction of the Corn–DFHO complex facilitates label-free detection of lncRNA with a higher signal-to-background ratio than the non-sequence-specific dye SYBR Gold. Taking advantage of the high efficiency of DSN/T7 transcription-mediated cascade amplification and high signal-to-background ratio of Corn–DFHO complexes, this Corn-based biosensor can sensitively detect lncRNA with a LOD of 31.98 aM.…”

Synergy of Target-Induced Magnetic Network and Single-Drop Chromogenic System for Ultrasensitive “All-in-Tube” Detection of miRNA in Whole Blood

“…In clinical settings, there has been increasing attention on the study of circulating tumor RNA (ctRNA, including microRNA) . MicroRNA (miRNA), a kind of noncoding RNA molecules, is widely present in eukaryotic cells and plays a crucial roles in gene regulation. The dysregulation of miRNA expression is intricately linked to the development and progression of various types of tumors. − Nevertheless, due to the features of low abundance, short nucleotide sequences, and high similarity among family members, conventional detection methods face challenges in analyzing miRNA expression profiles .…”

Efficient Three-Dimensional DNA Nanomachine Guided by a Robust Tetrahedral DNA Nanoarray Structure for the Rapid and Ultrasensitive Electrochemical Detection of Matrix Metalloproteinase 2

“…On account of these drawbacks, three-dimensional (3D) DNA nanomachines immobilized on the surface of AuNPs and magnetic beads (MBs) were proposed. , Benefiting from their high DNA-loading capability, they definitely improved the walking speed and signal amplification efficiency to some extent, but some remaining problems still cannot be ignored. The majority of the traditional 3D DNA nanomachines not only moved on the disordered DNA tracks composed of two-dimensional (2D) or one-dimensional (1D) single-stranded (ss)­DNA probes, , but also walked in a ″step-by-step″ fashion with a limited number and length of arms, , which readily led to a deviation from the designed tracks so as to greatly limit the amplification efficiency and slow down the walking rate of the 3D DNA nanomachines. Surprisingly, it was well known that the tetrahedral DNA (TDN) structure has the merits of a predesigned size, structural stability, and excellent biocompatibility. − Hence, it was prospective to make the 3D DNA nanomachine walk with better directionality and a faster reaction rate by using TDNs as rolling and amplification scaffolds.…”