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- Titre iSpinach: an optimized fluorogenic RNA aptamer
- Description The recent development of new fluorogenic dyes and the concomitant isolation of specific RNA aptamers offer the possibility to monitor in real-time the synthesis of a specific RNA both in vitro and in vivo. Among this new generation of dyes, the 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI), a commercially available dye mimicking the natural fluorophore of the green fluorescent protein, proved to be particularly well suited for live-cell imaging as it is nontoxic, cell membrane-permeable, does not interact with cell components and has a low fluorescence in it is free state. These attractive properties led to the isolation of a first DFHBI-binding aptamer termed Spinach. While Spinach was able to enhance DFHBI fluorescence more than ~2000 times upon binding, it however suffered from several limitations such as limited folding efficiency and thermal instability. These limitations were partly overcome in a second version of the aptamer (Spinach2) obtained by rational design.
- Bénéfices iSpinach has been developed to circumvent important limitations of the existing DFHBI-binding aptamer:
– Efficient folding in potassium-free environment
– High folding efficiency
– High thermal stability
– Improved brightness
– 1.3 times shorter than Spinach2
- Nouveauté This invention provides an improved version of the RNA aptamer Spinach = iSpinach, a nucleic acid molecule capable of binding to a fluorophore molecule, in particular DFHBI. iSpinach is not only able to interact with the fluorophore DFHBI in the presence of potassium to form a complex more fluorescent but it can also do it in the absence of any monovalent cation and at physiological temperature.
- Mots clés RNA, Entamer, Fluorogenic dyes, Cell imaging, HTS
- Secteurs Health
