Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology

Area of Science:

• Molecular Biology
• Biochemistry
• Synthetic Biology

Background:

• RNA aptamers offer modular fluorescence visualization of native RNAs.
• Split aptamers enable real-time tracking of RNA assemblies with low background noise.
• Split aptamers eliminate the need for covalently attached dyes.

Purpose of the Study:

• To design and test F30 Broccoli aptamer splits (fluorets) for fluorescence and stability.
• To establish methods for assembling split aptamers for cellular applications.
• To develop rules for constructing responsive biomaterials and programmable fluorescent systems.

Main Methods:

• Design and characterization of F30 Broccoli aptamer splits (fluorets).
• Assembly of split aptamers via one-pot thermal annealing and co-transcription.
• Testing of responsive biomaterials with stimuli (divalent ions, nucleases, temperature).
• Application of strand displacement for isothermal fluorescent response control.

Main Results:

• Demonstrated successful assembly of split aptamers through annealing and co-transcription.
• Showcased direct tracking of transcription reactions using fluorescent output.
• Established rules for creating stimuli-responsive fluorescent biomaterials.
• Validated strand displacement for programmable isothermal fluorescence control.

Conclusions:

• Split RNA aptamers (fluorets) provide a versatile platform for dynamic RNA visualization and monitoring.
• This work enables the development of responsive biomaterials and molecular computing systems.
• The findings lay the groundwork for biocompatible logic gates and real-time cellular process monitoring.