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Simple preparation of biotinylated RNA by transcription via an unnatural base pair.

To develop a convenient method for RNA fragment biotinylation at a specific position, we synthesized ribonucleoside 5'-triphosphates of biotinylated unnatural bases. These unnatural bases were composed of 2-oxo(1H)pyridine (denoted as y), specifically pairing with 2-amino-6-(2-thienyl)purine (denoted as s) in transcription. The y base was linked with biotin through an allylamine or propynylamine linker at position 5 of y. These triphosphate derivatives were specifically incorporated by T7 RNA polymerase into RNA opposite s in templates. The specific biotinylation and immobilization of an RNA aptamer were demonstrated by this system.