Isolation and Identification of Post-Transcriptional Gene Silencing-Related Micro-RNAs by Functionalized Silicon Nanowire Field-effect Transistor

Abstract

Many transcribed RNAs are non-coding RNAs, including microRNAs (miRNAs), which bind to complementary sequences on messenger RNAs to regulate the translation efficacy. Therefore, identifying the miRNAs expressed in cells/organisms aids in understanding genetic control in cells/organisms. In this report, we determined the binding of oligonucleotides to a receptor-modified silicon nanowire field-effect transistor (SiNW-FET) by monitoring the changes in conductance of the SiNW-FET. We first modified a SiNW-FET with a DNA probe to directly and selectively detect the complementary miRNA in cell lysates. This SiNW-FET device has 7-fold higher sensitivity than reverse transcription-quantitative polymerase chain reaction in detecting the corresponding miRNA. Next, we anchored viral p19 proteins, which bind the double-strand small RNAs (ds-sRNAs), on the SiNW-FET. By perfusing the device with synthesized ds-sRNAs of different pairing statuses, the dissociation constants revealed that the nucleotides at the 3′-overhangs and pairings at the terminus are important for the interactions. After perfusing the total RNA mixture extracted from Nicotiana benthamiana across the device, this device could enrich the ds-sRNAs for sequence analysis. Finally, this bionanoelectronic SiNW-FET, which is able to isolate and identify the interacting protein-RNA, adds an additional tool in genomic technology for the future study of direct biomolecular interactions.

Application of Silicon Nanowire Field Effect Transistor (SiNW-FET) Biosensor with High Sensitivity

“…In other works in the literature, 3-mercapto propyl trimethoxysilane (MPTMS) has been proposed to form a thiol (SH)-termination to modify the surface. Chen et al [ 37 ] were able to modify DNA probes using a disulfide bond formed on the SiNW surface by MPTMS. 3-glycidyloxy propyltrimethoxysilane (GPTES) is a kind of epoxy silane that can form a homogeneous silane monolayer in buffer solutions (pH 8–9).…”

Silicon Nanowires and Their Impact on Cancer Detection and Monitoring

“…Previous works detecting miRNA using SiNW-FET have been documented 117 and have demonstrated a 7-fold higher sensitivity than rt-PCR. 118 Additionally, while significant progress has been made using SiNWs for the capture and enrichment of CTCs 94,95,99 and ECVs, 102 none of the previous work involved SiNWs in directly detecting elements of liquid biopsies, whether by electrical or optical detection. Vertically aligned SiNW arrays can be used to enrich liquid biopsy entities (Figure 5), combining this process with the previously discussed photoluminescence or electrical modes of detection of proteins and/ or DNA may provide crucial next steps in advancing the field of liquid biopsies toward direct and label-free detecting of liquid biopsy elements.…”

“…(D) miR21probe/SiNW-FET detecting RNA extracts from two different cancer cell lines detecting miR21 RNA. Parts B and C are adapted from the article “Isolation and Identification of Post-Transcriptional Gene Silencing-Related Micro-RNAs by Functionalized Silicon Nanowire Field-effect Transistor” (ref ).…”

“…Combining enrichment techniques for tumor derived ECVs discussed earlier (postenrichment processing to digest ECVs and harvest mi-RNA) with SiNW-FETs based sensing of miRNA could provide a valuable detection scheme for ct-miRNA unbound to proteins. Previous works detecting miRNA using SiNW-FET have been documented and have demonstrated a 7-fold higher sensitivity than rt-PCR …”

Structural insights into interactions between viral suppressor of RNA silencing protein p19 mutants and small RNAs

“…New strategies for detection, quantification and sequestration of miRNAs in living systems and in biological samples such as human serum have been critical for understanding miRNA function and modulating their activity towards human therapeutics. Because of its unique ability to bind small RNA duplexes of any sequence, p19 has been applied in a range of creative strategies for small RNA detection and sequestration [7,[14][15][16][17][18][19][20][21][22]. miRNA duplexes, before being unwound by RISC, are very similar to p19's canonical ligands, 21-nt vsiRNAs, except that they are 21-23 nt long and typically contain numerous non-Watson-Crick base pair mismatches in their sequence [23,24].…”