Isolation of a site-specifically modified RNA from an unmodified transcript

Abstract

Natural RNAs contain many base modifications that have specific biological functions. The ability to func-tionally dissect individual modifications is facilit-ated by the identification and cloning of enzymes responsible for these modifications, but is hindered by the difficulty of isolating site-specifically modified RNAs away from unmodified transcripts. Using the m1G37 and m1A58 methyl modifications of tRNA as two examples, we demonstrate that non-pairing base modifications protect RNAs against the DNA-directed RNase H cleavage. This provide a new approach to obtain homogeneous RNAs with site-specific base modifications that are suitable for biochemical and functional studies