Nowadays, RNA synthesis has become an essential tool not only in the field of molecular biology and medicine, but also in areas like molecular diagnostics and material sciences. Beyond synthetic RNAs for antisense, aptamer, ribozyme, and siRNA technologies, oligoribonucleotides carrying site-specific modifications for structure and function studies are needed. This often requires labeling of the RNA with a suitable spectroscopic reporter group. Herein, we describe the synthesis of functionalized monomer building blocks that upon incorporation in RNA allow for selective reaction with a specific reporter or functional entity. In particular, we report on the synthesis of 5′-O-dimethoxytrityl-2′-O-tert-butyldimethylsilyl protected 3′-O-phosphoramidites of nucleosides that carry amino linkers of different lengths and flexibility at the heterocyclic base, their incorporation in a variety of RNAs, and postsynthetic conjugation with fluorescent dyes and nitroxide spin labels. Further, we show the synthesis of a flavine mononucleotide-N-hydroxy-succinimidyl ester and its conjugation to amino functionalized RNA.

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SummaryOver the past 20 years, the generation of functional RNAs by in vitro selection has become a standard technique. Apart from aptamers for simple binding of defined ligands, also RNAs for catalysis of chemical reactions have been selected. In the latter case, a key step often is the conjugation of one of the two reactants to the library, requiring suitable strategies for terminal or internal RNA functionalization. With the aim of selecting a ribozyme for deamination of cytidine, we have set up a selection scheme involving the attachment of the cytidine acting as deamination substrate to the 3'-terminus of the RNAs in the library, and library immobilization. Here, we report the synthesis of a bifunctional cytidine derivative suitable for conjugation to RNA and linkage of the conjugated library to a streptavidine-coated surface. Successful conjugation of the cytidine derivative to the 3'-terminus of a model RNA is demonstrated.