Characterization of Hairpin Ribozyme Reactions

Hairpin ribozymes are small RNA catalytic motifs naturally found in the satellite RNAs of tobacco ringspot virus (TRsV), chicory yellow mottle virus (CYMoV), and arabis mosaic virus (ArMV). The catalytic activity of the hairpin ribozyme extends to both cleavage and ligation reactions. Here we describe methods for the kinetic analysis of the self-cleavage reaction under transcription reaction conditions. We also describe methods for the generation of DNA templates for subsequent in vitro transcription reaction of hairpin ribozymes. This is followed by a description of the preparation of the suitable RNA molecules for ligation reaction and their kinetic analysis.