Genome-wide analysis of Vibrio cholerae cytolysin (VCC) in T84 monolayer cells

Non-toxic cocentrations of VCC induces upregulation of inflammatory genes in T84 monolayer cells Stimulation with VCC was done from the apical side of polarized tight monolayers of T84 cells by replacing the LB medium in the upper chamber with 0.5 mL challenge medium containing 160 ng/ml (VCC). Experimental control was done with challenge LB medium only. The challenge was done under aerobic condition and at 37°C for 5 hours. Total RNA was extracted from T84 polarized tight monolayers by using the Qiagen RNeasy Mini Kit according to the manufacturer's instructions and dissolved in RNase-free water containing Promega rRNasin ribonuclease inhibitor. Quality control was performed with Agilent 2100 Bioanalyser and NanoDrop.