To identify candidate genes whose expressions were regulated by the histone H4 modifications during P. aeruginosa PA01 exposure
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ABSTRACT: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for the histone modifications H4K5ac and H4K8ac in mid-L4 WT worms exposed to P. aeruginosa PA01 for 12 hours.
Project description:To identify candidate genes whose expressions were regulated by P. aeruginosa PA01, we used RNA-Seq analysis to study the transcriptomic profiles of mid-L4 worms exposed to P. aeruginosa PA01 for 12 hours. Differential expression analysis was performed by comparing WT worms exposed to P. aeruginosa PA01 for 12 hours versus worms fed E. coli OP50.
Project description:Differential expression of regulatory genes of reference strain P. aeruginosa PA01, involved in the quorum sensing was analyzed using Microarray. Total RNA was isolated from reference strain P. aeruginosa PA01, grown with or without bacterial extracts (1.0 mg/ml) using TRI reagent. Total RNA was quantified, and 10 µg RNA was converted to cDNA, fragmented and labelled by following GeneChip P. aeruginosa PA01 genome array user manual. Labelled cDNAs were hybridized with P. aeruginosa genome array gene chip, washed and stained. Hybridized chips were scanned, processed and analyzed using expression console and transcriptome analysis console.
Project description:Differential expression of regulatory genes of reference strain P. aeruginosa PA01, involved in the quorum sensing was analyzed using Microarray. Total RNA was isolated from reference strain P. aeruginosa PA01, grown with or without bacterial extracts (0.1 mg/ml) using TRI reagent. Total RNA was quantified, and 10 µg RNA was converted to cDNA, fragmented and labelled by following GeneChip® P. aeruginosa PA01 genome array user manual . Labelled cDNAs were hybridized with P. aeruginosa genome array gene chip , washed and stained. Hybridized chips were scanned, processed and analyzed using expression console and transcriptome analysis console.
Project description:The transcriptome of P. aeruginosa PA01 biofilm cells was compared to the associated suspended culture, upon growth on reverse osmosis membrane coupon under limited nutrient conditions. Keywords: growth in reverse osmosis unit and on reverse osmosis membrane
Project description:ChIP-on-chip experiments with MvaT-V and MvaU-V (V = VSVG epitope tag) using a P. aeruginosa PA01 high-density oligonucleotide array designed and manufactured by Roche NimbleGen, Inc.
Project description:In the clinical setting, mutations in the CFTR gene enhance the inflammatory response to P. aeruginosa (PA01) infection, but measurements of the inflammatory response to pathogen stimulation by isolated airway epithelia can yield variable results. In this series, we exposed CFBE41o- cells over-expressing ?F508/?F508 CFTR and CFBE41o- cells rescued with wt-CFTR to P. aeruginosa biofilms. P. aeruginosa elicited a more robust increase in cytokine and chemokine expression (e.g., IL-8, CXCL2, CXCL3, CXCR4 and TNF-?) in CFBE-wt-CFTR cells compared to CFBE-?F508-CFTR cells. These results demonstrate that CFBE41o- cells complemented with wt-CFTR mount a more robust inflammatory response to P. aeruginosa than CFBE41o- ?F508/?F508-CFTR cells. CFBE41o- cells generously provided by Dr. J.P. Clancy (University of Alabama). The series involved 4 treatment groups: unexposed wild type CFTR cells, unexposed ?F508/?F508 CFTR cells, wild type CFTR cells exposed to PA01, and ?F508/?F508 CFTR cells exposed to PA01. Each treatment group involved 4 replicate polarized monolayers. PAO1 was added to the apical side of exposed monolayers at a multiplicity of infection (MOI) of 30:1 for 1 hour in the absence of antibiotics, and then planktonic PAO1 was removed by replacing the apical medium with MEM supplemented with 0.4% arginine (2). Control monolayers were treated identically except that vehicle only (medium used to grow PAO1, MEM supplemented with 0.4% arginine) was added to the apical side of cells. Five hours after washing planktonic P. aeruginosa from the cell monolayers mRNA was isolated.
Project description:In the clinical setting, mutations in the CFTR gene enhance the inflammatory response to P. aeruginosa (PA01) infection, but measurements of the inflammatory response to pathogen stimulation by isolated airway epithelia can yield variable results. In this series, we exposed CFBE41o- cells over-expressing ∆F508/∆F508 CFTR and CFBE41o- cells rescued with wt-CFTR to P. aeruginosa biofilms. P. aeruginosa elicited a more robust increase in cytokine and chemokine expression (e.g., IL-8, CXCL2, CXCL3, CXCR4 and TNF-α) in CFBE-wt-CFTR cells compared to CFBE-∆F508-CFTR cells. These results demonstrate that CFBE41o- cells complemented with wt-CFTR mount a more robust inflammatory response to P. aeruginosa than CFBE41o- ∆F508/∆F508-CFTR cells.
Project description:The transcriptome of P. aeruginosa PA01 biofilm cells was compared to the associated suspended culture, upon growth on reverse osmosis membrane coupon under limited nutrient conditions. Experiment Overall Design: Cells were collected from the RO unit and from the membrane coupon after 20 hours of growth and biofouling of the membrane. A defined media was used, resembeling secondary wastewaters.