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Real-time Imaging of Single Engineered RNA Transcripts in Living Cells Using Ratiometric Bimolecular Beacons
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Programmed Fluorescence-Encoding DNA Nanoflowers for Cell-Specific-Target Multiplexed MicroRNA Imaging.

Fan Zhang1, Wenhao Dai1, Meiqin Zhang1

  • 1Beijing Key Laboratory for Bioengineering and Sensing Technology, School of Chemistry and Biological Engineering, University of Science & Technology Beijing, Beijing 100083, P. R. China.

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Summary

This study introduces a novel DNA nanoflower system for multiplexed microRNA (miRNA) imaging in living cells. This innovative approach enables precise detection of multiple miRNAs for disease biomarker discovery.

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Nanotechnology

Background:

  • Accurate identification of multiple microRNAs (miRNAs) in specific cells is challenging due to limited fluorophores and lack of cell-specific targeting.
  • Existing methods struggle with high spatial resolution and multiplexed detection of intracellular miRNAs.

Purpose of the Study:

  • To develop a programmed fluorescence-encoding DNA nanoflower (CNFs) system for multiplexed miRNA imaging in living cells.
  • To enhance cellular targeting specificity and enable simultaneous detection of multiple intracellular miRNAs.

Main Methods:

  • Utilized rolling circle amplification (RCA) for self-assembly of CNFs.
  • Engineered CNFs with CD63 aptamer for cell targeting, dual fluorophore encoding for 9 distinct barcodes, and molecular beacons (MBs) for miRNA recognition.
  • Applied the CNFs system for evaluating nine miRNA expression profiles in breast cancer cells.

Main Results:

  • Successfully developed a CNFs system for multiplexed miRNA imaging in living cells.
  • Achieved enhanced cellular targeting and endocytic uptake via CD63 aptamer.
  • Demonstrated simultaneous detection of multiplexed intracellular miRNAs using CNFs conjugated with MBs.
  • Evaluated nine miRNA expression profiles in breast cancer, showcasing the system's utility.

Conclusions:

  • The developed CNFs system provides a valuable tool for multiplexed miRNA biomarker imaging in specific cells.
  • This technology facilitates the exploration of miRNA molecular regulation mechanisms in diseases like cancer.
  • The CNFs system offers a promising approach for advancing diagnostics and understanding miRNA functions.