Fig 1.

Deletion of rpl32 genes reduced ribosome levels in haploid cells. (A) Deletion of rpl32 paralogous genes in S. pombe using a PCR-based approach with long DNA tracts of flanking homology to the target locus. (B) Deletion mutants were confirmed by PCR with genomic DNA as a template and locus-specific primers flanking the target sites. (C) QPCR quantification of rpl32-1 and rpl32-2 transcript abundance in deletion mutants and WT cells cultured in YEPD medium. beta-actin was used to standardize transcript levels of paralog rpl32-1 and rpl32-2. (D) Western blot analysis of RPL32 in deletion mutants and WT cells cultured in YEPD medium. (E) Polysome profiles of deletion mutants and WT cells cultured to the late exponential phase (OD₆₀₀ of 5.0) in YEPD medium. A total of 15 A₂₆₀ units of extracts from each culture was analyzed on 5% to 50% sucrose gradients centrifuged for 4 h at 44,000 rpm (25).