Dye-Conjugated Spinach RNA by Genetic Alphabet Expansion.

Kyung Hyun Lee, Michiko Kimoto, Gota Kawai, Itaru Okamoto, Andrea Fin, Ichiro Hirao

Chemistry (Weinheim an der Bergstrasse, Germany) 2022 Mar 16

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Light-emitting systems using an RNA aptamer-dye pair, such as Spinach RNA, are an attractive method for imaging and tracing RNA expression in vitro and in vivo. We present an alternative Spinach method by genetic alphabet expansion using an unnatural base pair system, in which a dye-conjugated unnatural base substrate is site-specifically incorporated at a specific position in Spinach RNA by transcription involving the third base pair. The incorporation position was predicted by molecular dynamics simulations. This dye-conjugated Spinach RNA increased the thermal stability of the fluorescence, the robustness against ion sensitivity, and the resistance against photobleaching. Furthermore, we applied our method to Baby Spinach, a shorter version of Spinach, for dye conjugation toward the visible detection of transcripts. This is the first demonstration of an alternative RNA imaging method for a detection system using genetic alphabet expansion. © 2022 Wiley-VCH GmbH.

Citation

Kyung Hyun Lee, Michiko Kimoto, Gota Kawai, Itaru Okamoto, Andrea Fin, Ichiro Hirao. Dye-Conjugated Spinach RNA by Genetic Alphabet Expansion. Chemistry (Weinheim an der Bergstrasse, Germany). 2022 Mar 16;28(16):e202104396