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Studying Single mRNA Molecules from Cradle to Grave
Summary: Rob Singer’s lab seeks to understand the expression and movement of mRNA. His lab developed methods to label RNA endogenously in cell lines and mice. He develops microscopy to observe and quantify single mRNAs in neuronal processes, and can follow the mRNA from transcription through nuclear export, translation and degradation.
We seek to understand the expression and movement of mRNA from transcription through degradation and the effect that defects in these processes have on the cell. We develop methods to label RNA in fixed and living cells using fluorescent probes and microscopy techniques and image analysis algorithms to visualize and quantify many mRNAs simultaneously. Using these technologies, we can observe single mRNAs as they are transcribed and processed, as they export from the nucleus, localize to cytoplasmic compartments such as dendritic processes of neurons, and finally when and where they translate and are degraded. Because these techniques yield quantitative fluorescence data, we are able to apply mathematical modeling to test mechanistic hypotheses about the kinetics of each of the events in an mRNA’s life. Most recently, we have developed techniques which allow is to follow the mRNAs in cells in their native habitat; in tissues of the living animal. By making transgenic, knock-in mice where the endogenous RNA is tagged with the stem loops of the MS2 phage, and bound to the fluorescent coat protein, we can see all these events of mRNA from birth to death in living tissues. Our goal at Janelia is to develop microscopic approaches to observe these events in animals such as mice, worms and flies. We feel that this is the only way to ultimately understand the regulatory mechanisms governing gene expression.
As of September 06, 2012
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