Analysis of snoRNAs in wild-type, rnt1Δ, tgs1Δ, and rnt1Δ tgs1Δ strains. (A) Growth of wild-type, rnt1Δ, tgs1Δ, and rnt1Δ tgs1Δ strains. Shown are sister spores obtained after dissection of a tetrad. Spores were streaked on YPD medium and grown at 23°C for 6 days. (B) Analysis of the 5′ end of the snR50, snR64, and snR69 snoRNAs by primer extension in wild-type, rnt1Δ, tgs1Δ, and rnt1Δ tgs1Δ strains. Legend as in Figure 2. (C) Immunoprecipitation analysis of the TMG cap status of precursors and mature snoRNAs in wild-type, rnt1Δ, xrn1Δ rat1-1, and rnt1Δ tgs1Δ strains using the K121 and RR131 antibodies. Total RNAs, RNAs selected after binding to protein G beads coupled to K121 anti-TMG antibodies or to 12CA5 (anti-HA) antibodies, and RNAs selected after binding to protein A beads coupled to polyclonal R1131 anti-TMG antibodies or anti-GST antibodies were loaded on a 6% polyacrylamide sequencing gel, transferred to a Hybond N+ membrane, and probed with snR64-Rev and snR69-Rev antisense oligonucleotide probes hybridizing to the indicated snoRNAs.
