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Tandem Spinach Array for mRNA Imaging in Living Bacterial Cells
Jichuan Zhang1,2, Jingyi Fei1, Benjamin J Leslie1,3
1Department of Physics and Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, Urbana, IL 61801 USA.
Scientific Reports
|November 28, 2015
Summary
Researchers improved live cell RNA imaging by creating tandem arrays of Spinach aptamers. These arrays significantly boost mRNA brightness and do not disrupt RNA processes, enabling better visualization of RNA in cells.
Area of Science:
- Molecular Biology
- Biophysics
- Cell Biology
Background:
- Live cell RNA imaging is crucial for understanding RNA dynamics.
- Current Spinach aptamer-fluorogen systems offer limited brightness for mRNA imaging.
- Potential perturbations of RNA characteristics by aptamer-fluorogen systems are not fully understood.
Purpose of the Study:
- To enhance the brightness of aptamer-fluorogen systems for mRNA imaging.
- To assess the impact of aptamer arrays on native RNA transcription, translation, and degradation.
- To develop a generalizable labeling method for high-performance, low-perturbation live cell RNA imaging.
Main Methods:
- Constructed and tested tandem arrays of Spinach aptamers (8-64 repeats).
- Quantified brightness enhancement of tagged mRNA molecules in living cells.
- Evaluated mRNA transcription, translation, and degradation rates in cells with transcriptional fusions.
Main Results:
- Tandem Spinach aptamer arrays increased tagged mRNA brightness by up to ~17 fold.
- Pulsed laser excitation further improved imaging sensitivity of Spinach array-tagged RNAs.
- Transcriptional fusion of Spinach arrays did not significantly alter mRNA transcription, translation, or degradation.
Conclusions:
- Tandem Spinach aptamer arrays provide a significant brightness enhancement for live cell mRNA imaging.
- This labeling method demonstrates low perturbation of native RNA biological processes.
- Aptamer arrays represent a promising, generalizable strategy for advanced live cell RNA visualization.
