Ribo-Pop has minimal effects on expression measurements of non-rRNA genes. RNA-seq libraries prepared after Ribo-Pop depletion were compared to libraries made from nondepleted input RNA. To allow sufficient coverage for analysis, libraries were prepared using a poly(A)-primed method (QuantSeq). (A) Scatterplot and the Pearson's correlation between input and depleted RNA levels for mRNAs and ncRNAs. The RNA levels are the averages of three replicates for each condition, filtered to include only genes with an average of at least 1 count per million (CPM). Each replicate was prepared from a different RNA sample and corresponds to the same material used for the non-poly(A)-primed libraries analyzed in Figure 4. (B) MA plot for the comparison of Ribo-Pop depleted versus input libraries analyzed with DESeq2 (Love et al. 2014). Genes with an adjusted P-value <0.01 for differential expression are highlighted in pink if increased upon depletion or blue if decreased upon depletion. The two rRNA pseduogenes that are among the changing genes are indicated. (C) Stretches of complementarity between the Ribo-Pop probes and nontarget RNAs were identified by aligning the probe target sites to the transcriptome with BLAST (Altschul et al. 1990). Alignments are classified as overlapping a gene decreased upon Ribo-Pop depletion (decreased) or overlapping a gene not decreased upon depletion (other). Only alignments with 100% identity are displayed.
