FIGURE 3.

Survey of 5′-adenylation with 17-mer RNA substrate S1 and various lengths of DNA blocking oligo relative to the RNA 5′-end. Either no blocking oligo was included (far left set of lanes), or the blocking oligo n/0 ranged from n = 0 to +6 (i.e., 0–6 nt RNA 5′-overhang). Gel images from two experiments are shown; the +4/0 experiment is included on both gel images to facilitate comparison. For all sets of lanes, the four time points are at 0.5 min, 10 min, 1 h, and 6 h (20% PAGE). The various species are labeled as follows, bottom to top: circ, circularization; un, unreacted RNA; App, 5′-adenylation; lig, blocking oligo ligation; olig, oligomerization (see Fig. 2 ▶). Similar to the 0/0 experiment (i.e., n = 0), no reaction was observed with blocking oligos for which n is negative (i.e., a recessed RNA 5′-end; data not shown).