Figure 8. Characterization of S. pombe Safs.

A) Protein G pull-downs from the indicated S. pombe strains were blotted with antibodies to the HA epitope. The bands with asterisks correspond to the indicated proteins and were quantified relative to background. B and C) Spores from the (B) saf2::ura4⁺/saf2⁺ and (D) saf3::ura4⁺/saf3⁺ diploids were germinated in minimal medium lacking uracil. Cells were fixed in formaldehyde at 15 and 40 h, respectively, and stained with DAPI. D and E) RNA was purified from wildtype cells grown at 32°C, prp2-1 cells grown at 25°C(−) or shifted to 36°C (+) for 4 hours, or from spores germinated at 32°C for 24 h from saf2::ura4⁺/saf2⁺ (D) and saf3::ura4⁺/saf3⁺ (E) diploids in medium lacking uracil. RT-PCR reactions were performed using oligonucleotides that flank the long intron within the prp19 mRNA. PCR products were separated on 3% Nusieve gels and detected with ethidium-bromide. Arrows indicate the position of prescursor and mature RNA species.