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Fluorophore‐binding RNA aptamers and their applications

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Why image RNA? Of all the biological molecules, RNA exhibits the most diverse range of functions. Evidence suggests that transcription produces a wide range of noncoding RNAs (ncRNAs), both short (e.g., siRNAs, miRNAs) and long (e.g., telomeric RNAs) that regulate many aspects of gene expression, including the epigenetic processes that underlie cell fate determination, polarization, and morphogenesis. All these functions are realized through the exquisite temporal and spatial control of RNA expression levels and the stability of specific RNAs within well‐defined sub‐cellular compartments. Given the central importance of RNA in dictating cell behavior via gene‐related functions, there is a great demand for RNA imaging methods so as to determine the composition of the cellular ‘transcriptome’ and to acquire a complete spatial‐temporal profile of RNA localization. Recent advances in fluorophore‐binding RNA aptamers promise to provide exactly this knowledge, which can ultimately advance our understanding of cell function and behavior in conditions of health and disease, and in response to external stimuli. WIREs RNA 2016, 7:843–851. doi: 10.1002/wrna.1383 This article is categorized under: RNA Interactions with Proteins and Other Molecules > Small Molecule–RNA Interactions RNA Methods > RNA Analyses In Vitro and In Silico RNA Methods > RNA Analyses in Cells
The effects of KD and FE on RNA imaging contrast. Comparison of MS2 (a, b), RNA Spinach (c), and RNA Mango (d) systems. Figure was generated assuming that free fluorophore in each example is at a fixed multiple of the KD. (a) A single repeat of the MS2 RNA tag has high binding, but poor contrast. (b) Multiple copies of the MS2 tag (here 6 are shown, typically more are used) increases contrast by enhancing FE. (c) RNA Spinach, while having an excellent FE, requires high levels of free fluorophore, which lowers contrast. (d) RNA Mango can operate with comparable brightness to the Spinach system but at much lower free fluorophore concentration resulting in a potentially higher contrast. Circles represent a field of view. Asterisks symbolize the fluorophore with size reflecting brightness. In the MS2 system (a, b) the fluorophore is always ‘ON.’ The background color represents the averaged signal generated by the unbound fluorophore in each system.
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RNA Interactions with Proteins and Other Molecules > Small Molecule–RNA Interactions
RNA Methods > RNA Analyses In Vitro and In Silico
RNA Methods > RNA Analyses in Cells

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