The purpose of this proposal is to build on our development of single molecule RNA detection in living cells in real time. 1. To extend the technology for fast 4D imaging to describe the movements of RNA transcripts in the nucleus and cytoplasm. 2. To develop a method for detecting translation of single RNA molecules. To image RNA transcription in living cells and follow the individual transcripts released using high speed microscopic 4D imaging to correlate the molecular steps of processing, export and localization with the spatial and temporal colocalization with specific proteins. This work will result in the ability to define the mechanics of RNA metabolism (nuclear export, cytoplasmic transport, localization, translation and degradation) important for the normal functioning of all eukaryotic cells within a spatial and temporal cellular context heretofore not possible using current methods. ? ?
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