Illumina HiSeq 1500 paired end sequencing; RNA-seq of coding RNA of Staphylococcus aureus treated with the antimicrobial coating AgXX reveals strong thiol-specific stress response and protein S-bacillithiolation in Staphylococcus aureus
RNA-seq of coding RNA of Staphylococcus aureus treated with the antimicrobial coating AgXX reveals strong thiol-specific stress response and protein S-bacillithiolation in Staphylococcus aureus
Strain cultivation was performed in RPMI1640  medium (Lonza) under vigorous agitation at 37°C in triplicate until cells have reached an optical density at 500 nm of 0.5. S. aureus cells were harvested before and 30 min after exposure to 4.5 µg/ml AgXX373 Strain cultivation was performed in RPMI1640  medium (Lonza) under vigorous agitation at 37°C in triplicate until cells have reached an optical density at 500 nm of 0.5. S. aureus cells were harvested before and 30 min after exposure to 4.5 µg/ml AgXX373. S. aureus cells with an optical density at 500 nm of 0.5 were treated 30 min 4.5 µg/ml AgXX373 RNA isolation was performed using the phenol-chloroform-isoamylalcohol approach. After precipitation with 3 M sodium acetate and isopropanol, the total RNA was washed with cold ethanol and the pellet was solved in sterile water. Initially RNA quality was checked by Trinean Xpose (Gentbrugge,Belgium) and Agilent RNA Nano 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). Samples contaminated with DNA were treated with DNase (Qiagen), cleaned as described above and rechecked by Xpose and Agilent Bioanalyzer. Finally RNA was free of DNA with an RNA Integrity Number (RIN) > 9 and rRNA Ratio [23s / 16s] > 1.5 Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina (San Diego, CA, USA) was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was checked by Agilent RNA Pico 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). RNA was free of detectable rRNA. TruSeq Stranded mRNA Library Prep Kit from Illumina (San Diego, CA, USA) was used to prepare cDNA libraries.
Freie Universitat Berlin Institut fur Biologie-Mikrobiologie
E-MTAB-7074:USA300_AgXX_1_strRNA_B4_p
Experimental Factor: compound
AgXX373
Experimental Factor: dose
4.5