FIGURE 1.

Outline of the ligation method to study RNA modifications. (A) The RNA (black line) containing an unmodified or modified nucleotide (open or filled black circles) is hybridized with two oligonucleotides. These oligonucleotides are ligated by the T4 DNA ligase using this RNA as a template (triangle). The D- and N-oligos have different nucleotide compositions (green or red circles) at the ligation junction or different ligation sites. (B) For the D-oligo (green line), ligation proceeds very efficiently when the RNA is unmodified, but very poorly when the RNA is modified (or the other way around). For the N-oligo (red line), ligation has a constant yield regardless of the status of RNA modification. The amount of D-oligo ligation product corresponds to the amount of unmodified over modified RNA template (or modified over unmodified). The amount of N-oligo product corresponds to the sum of the unmodified and modified RNA.