Multicomponent DNA Nanomachines for Amplification-Free Viral RNA Detection
Abstract
1. Introduction
2. Results
2.1. Selection of RNA Fragments
2.2. Design of DNA Nanosensors
2.3. Detection by MB-DNSs
2.3.1. Estimation of LOD on a Synthetic Analyte
2.3.2. RT-PCR Amplicon Detection Results
2.3.3. Clinical Sample Detection Results
2.4. Detection by DNMs
2.4.1. DNM Assembly Visualization
2.4.2. Estimation of LOD for Synthetic DNA
2.4.3. Detection of Total Genomic RNA
2.4.4. Detection of RNA Extracted from Clinical Samples
3. Discussion
4. Materials and Methods
4.1. Materials
4.1.1. Detection Assay
4.1.2. RNA Extraction
4.1.3. RT-PCR
4.1.4. Electrophoresis
4.1.5. Purification of PCR Products
4.2. Methods
4.2.1. Objects of the Research
4.2.2. Design of MB-DNSs and DNMs
4.2.3. DNA Nanomachine Assembly
4.2.4. Polyacrylamide Gel Electrophoresis
4.2.5. RT-PCR
4.2.6. Agarose Gel Electrophoresis
4.2.7. Purification of Double-Stranded DNA Fragments
4.2.8. Fluorescent Detection Method: Detection by MB-DNSs
4.2.9. Fluorescent Detection Method: Detection by Multicomponent DNMs
4.2.10. Limit of Detection
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Criteria | MB-Based DNA Nanosensors | Deoxyribozyme DNA Nanomachines | RT-qPCR |
---|---|---|---|
Detection speed | 15–30 min | 30–180 min | 90–120 min |
Ease of development | Computer tools are used for designing Based on the principle of complementarity | Computer tools are used for designing Based on the principle of complementarity | Computer tools are used to select primers based on the principle of complementarity The amplification and visualization protocols are additionally selected, often empirically |
Sensitivity | Approximately 1–3 nM | Approximately 5–10 pM | Approximately 3 attoM |
Specificity | Specific | Specific | Specific |
Complexity | Does not require assembly step, easy to design | The assembly is necessary, the reaction is simple, require addition equipment to incubate samples, easy to design | Method requires specific expensive equipment, reagents and enzymes, laboratory condition, highly qualified personnel |
Cost of the analysis | USD 0.35 per reaction | USD 0.6 per reaction | USD 1.7 per reaction |
Competitiveness with existing methods | Cannot achieve the same sensitivity as PCR; works in combination as an alternative to visualization and recognition of specific PCR product | Cannot achieve the same sensitivity as PCR but able to achieve amplification-free detection of fM concentrations of ssRNA | Remains the gold standard for accurate quantification of nucleic acids. The success of detection strongly depends on the stages of sample preparation and amplification |
Feasibility of viral RNA detection | Effectively detects short ssDNA, PCR products | Detects short DNA analyte as well as long viral RNA samples | Detects DNA/RNA after amplification |
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© 2025 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
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Solyanikova, V.V.; Gorbenko, D.A.; Zryacheva, V.V.; Shtro, A.A.; Rubel, M.S. Multicomponent DNA Nanomachines for Amplification-Free Viral RNA Detection. Int. J. Mol. Sci. 2025, 26, 3652. https://doi.org/10.3390/ijms26083652
Solyanikova VV, Gorbenko DA, Zryacheva VV, Shtro AA, Rubel MS. Multicomponent DNA Nanomachines for Amplification-Free Viral RNA Detection. International Journal of Molecular Sciences. 2025; 26(8):3652. https://doi.org/10.3390/ijms26083652
Chicago/Turabian StyleSolyanikova, Valeria V., Daria A. Gorbenko, Valeriya V. Zryacheva, Anna A. Shtro, and Maria S. Rubel. 2025. "Multicomponent DNA Nanomachines for Amplification-Free Viral RNA Detection" International Journal of Molecular Sciences 26, no. 8: 3652. https://doi.org/10.3390/ijms26083652
APA StyleSolyanikova, V. V., Gorbenko, D. A., Zryacheva, V. V., Shtro, A. A., & Rubel, M. S. (2025). Multicomponent DNA Nanomachines for Amplification-Free Viral RNA Detection. International Journal of Molecular Sciences, 26(8), 3652. https://doi.org/10.3390/ijms26083652