RNA-seq of coding RNA of Streptococcus pneumoniae D39 after FCS addition
Streptococcus pneumoniae (S.p.) is the most common causative agent of community-acquired pneumonia worldwide. A key pathogenic mechanism that exacerbates severity of disease is the disruption of the alveolo-capillary barrier. However, the specific virulence mechanisms responsible for this in the human lung are not yet fully understood. RNA sequencing of Streptococcus pneumoniae transcriptome under infection media conditions, but without the presence of lung tissue, representing anon-host-infection scenario FCS+/- was analyzed,. RNA isolation was performed using an acidic phenol-chloroform extraction protocol (Wetzstein et al., 1992). After DNase-I treatment (Zymo Research, Germany), the RNA quality was checked by Trinean Xpose (Gentbrugge, Belgium) and the Agilent RNA Nano 6000 kit using an Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). For RNA-seq transcriptomics, Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina (San Diego, CA, USA) was used to remove the rRNA. TruSeq Stranded mRNA Library Prep Kit from Illumina (San Diego, CA, United States) was applied to prepare the cDNA libraries. The cDNAs were sequenced paired end on an Illumina HiSeq 1500 (San Diego, CA, United States) using 70 and 75?bp read length and a minimum sequencing depth of 10 million reads per library.