Fig. 3
Validation of dual RNA-seq. a We confirmed dual RNA-seq gene expression values by qRT-PCR. The infection study was repeated in duplicates and total RNA was isolated as previously described. Ten human and 19 pneumococcal genes were chosen as validation targets. We plotted fold changes from qRT-PCR against dual RNA-seq fold changes and observed a high degree of correlation for both species (R 2 > 0.7, Pearson). b We also confirmed pneumococcal gene expression at the protein level by quantitative fluorescence microscopy. Four target genes (SPD_0475, SPD_0963, SPD_1711, and SPD_1716) were C-terminally tagged with green fluorescent protein (GFP) at their own locus. GFP fusions were introduced in the ∆cps2E strain expressing red fluorescent protein (RFP) fused to HlpA. c Non-deconvolved image of SPD_1711-GFP up to 120 mpi. While RFP emitted a relatively constant signal, the GFP signal increased. d Dual RNA-seq expression values superimposed on the GFP/RFP ratio. To some extent, transcriptional changes corresponded to protein expression