Fluorescent and Bioluminescent Proteins Engineered to Visualize Biological Function in Single Living Cells

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June 1, 2011

A current focus of biological research is to quantify and image cellular processes in single living cells. To detect such cellular processes, genetically-encoded reporters have been extensively used. The most common reporters are firefly luciferase, renilla luciferase, green fluorescent protein (GFP) and its variants with various spectral properties. Herein, novel design of split GFP and split luciferase will be described; the principle is based on reconstitution of the split-reporter fragments when they are brought sufficiently close together. To demonstrate the usefulness of the split-GFP reporters, we have applied the reporters for developing a genetic method to identify mitochondrial proteins and their localization, and imaging dynamics of endogenous mRNA in single living cells. We have used a split-luciferase reporter for noninvasive imaging of protein-protein interactions in living cells and embryos. We have developed another design of reporter proteins; a cyclic luciferase by protein splicing to monitor protease activities in living cells. In this symposium, I will focus on the RNA imaging, different protein interactions and caspase activities with fluorescent and bioluminescent proteins in single living cells.

Molecular Biology

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