Debranchase-resistant labeling of RNA using the 10DM24 deoxyribozyme and fluorescent modified nucleotides

Abstract

The 10DM24 deoxyribozyme can site-specifically label RNAs with fluorophore-GTP conjugates; however, the 2′,5′-branched RNA linkage is readily cleaved by debranchase. To prevent loss of labels upon cleavage, we synthesized phosphorothioate-modified, fluorescent GTP derivatives and elaborated conditions for their incorporation by 10DM24. RNAs labeled with fluorescent derivatives of Sp-GTP^S were found to be resistant to debranchase.