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In Situ Spatial Complementation of Aptamer-Mediated Recognition Enables Live-Cell Imaging of Native RNA Transcripts in Real Time.

  • Author(s): Wang, Zejun
  • Luo, Yao
  • Xie, Xiaodong
  • Hu, Xingjie
  • Song, Haiyun
  • Zhao, Yun
  • Shi, Jiye
  • Wang, Lihua
  • Glinsky, Gennadi
  • Chen, Nan
  • Lal, Ratnesh
  • Fan, Chunhai
  • et al.
Abstract

Direct cellular imaging of the localization and dynamics of biomolecules helps to understand their function and reveals novel mechanisms at the single-cell resolution. In contrast to routine fluorescent-protein-based protein imaging, technology for RNA imaging remains less well explored because of the lack of enabling technology. Herein, we report the development of an aptamer-initiated fluorescence complementation (AiFC) method for RNA imaging by engineering a green fluorescence protein (GFP)-mimicking turn-on RNA aptamer, Broccoli, into two split fragments that could tandemly bind to target mRNA. When genetically encoded in cells, endogenous mRNA molecules recruited Split-Broccoli and brought the two fragments into spatial proximity, which formed a fluorophore-binding site in situ and turned on fluorescence. Significantly, we demonstrated the use of AiFC for high-contrast and real-time imaging of endogenous RNA molecules in living mammalian cells. We envision wide application and practical utility of this enabling technology to in vivo single-cell visualization and mechanistic analysis of macromolecular interactions.

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