FIG. 4.

Site-directed mutagenesis of snoRNA-GFP constructs. (A) Sequences of a portion of snR13 or snR47 3′ downstream RNAs. Mutations are identified above the sequences. GUA[AG] and UCUU motifs are highlighted. Bars below the sequences represent mutants identified in the selection experiment (Fig. 3) that greatly impair the elements and result in growth at high concentrations of 3AT. (B) Northern blot analysis of total RNA derived from NRD1-HA WT yeast transformed with single mutant plasmids. The blots were probed for GFP as well as an SCR1 loading control. The snR47-Δ70 and snR13-Δ4 constructs were set at a value of 1, and all other deletions were expressed as a fraction of these maxima. snR13-108 and snR47-204 constructs represent basal levels of GFP expression, and all mutations were made from these plasmids.