Comment[ArrayExpressAccession]	E-GEOD-29946
Public Release Date	2012-05-20						
Investigation Title	Genome wide transcriptional analysis of P. aeruginosa PAO1 response to kappa-opioid U-50,488 in poor nutrient medium						
Comment[Submitted Name]	Genome wide transcriptional analysis of P. aeruginosa PAO1 response to kappa-opioid U-50,488 in poor nutrient medium
Experiment Description	Here we examined virulence activation in Pseudomonas aeruginosa in response to the synthetic kappa opioid agonist U-50, 488 in nutrient poor media where growth conditions are limited and density dependent quorum sensing is not activated. We used a microarray to define the virulence-related genes in P. aeruginosa grown in liquid poor nutrient medium to determine the effect of kappa-opioid receptor agonist on virulence/lethal phenotype All biological samples for gene expression analysis were prepared in triplicate. P. aeruginosa MPAO1 cells collected from liquid culture at 7 hrs of growth in 1.) 0.1xTY medium containing tryptone 1 g/L and yeast extract, 0.5 g/L, or 2.) 0.1xTY+25 mM potassium phosphate buffer, pH 6.0, or 3.) 0.1xTY+200 uM U-50,488, or 4.) 0.1xTY+  25 mM potassium phosphate buffer, pH 6.0+ 200 uM U-50,488 were used for RNA isolation. Microarray analysis was performed using Affymetrix P. aeruginosa GeneChips (Affymetrix, Santa Clara, CA) at the University of Chicago Functional Genomics Facility						
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Term Source Name	EFO
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Term Source File	http://www.ebi.ac.uk/efo/efo.owl						
Person Last Name	ZABORINA	Alverdy	Zaborina				
Person First Name	Olga	John	Olga				
Person Mid Initials	Y						
Person Email	ozaborin@surgery.bsd.uchicago.edu						
Person Affiliation	University of Chicago						
Person Phone	773-702-3739						
Person Fax	773-834-4546						
Person Address	Surgery, University of Chicago, 5841 S Maryland, Chicago, IL, USA						
Person Roles	submitter						
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Protocol Name	P-GSE29946-1	P-GSE29946-6	P-GSE29946-5	P-GSE29946-3	P-GSE29946-2	P-GSE29946-4	P-GSE29946-7
Protocol Description	ID_REF = <br>VALUE = MAS5 normalized signal<br>Detection = <br>Detection p-value = 	After hybridization, arrays were washed and stained with Streptavidin Phycoerythrin. The fluorescent signals were amplified using a biotinylated antistreptavidin antibody in a Affymetrix GeneChip Fluidics Station (Affymetrix, Inc.) and the signals were detected in a GeneChip Scanner 3000 7G. The expression data were extracted using the Gene Chip Operating Software (MicroArray Suite 5.0 software, Affymetrix).	1.5 µg of fragmented and biotin labeled targets was hybridized to array for 16 h at 50°C and 60 rpm in an Affymetrix Hybridization Oven 640.	Cells were suspended in RNA protector solution (Bio-Rad), centrifuged 5,000 rpm, 10 min, 4C, lyzed in lysozyme TE solution followed by TRIzol. Total RNA was separated using PLG Phase lock gel followed by precipitation with isopropanol. DNA was removed with DNA-free kit	P. aeruginosa PAO1 grew in 0.1xTY supplemented when needed with 25 mM potassium phosphate buffer pH 6.0 hrs and/or 200 uM U-50,488, 7 hrs, 37C	Ten μg of each total RNA was processed for biotin labeled target preparation for hybridization to Pseudomonas aeruginosa expression arrays according to Genechip Expression analysis technical manual (Affymetrix, Inc).	Data were normalized data using RMA backround correction and Quantile normaliztion technique. The cel file were used for data analysis using ANOVA to identify differentially expressed genes [P-value <0.01 , Fold change >1.25]. dChip was used for analysis.
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Protocol Type	bioassay_data_transformation	image_aquisition	hybridization	nucleic_acid_extraction	grow	labeling	feature_extraction
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Publication Title	The use of fluoroquinolones in chronic otitis suppurativa.						
Publication Author List	van de Heyning PH, Pattyn SR, Valcke HD, van Caekenberghe DL, Claes J						
PubMed ID	3325935						
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Comment[SecondaryAccession]	GSE29946						
Comment[GEOLastUpdateDate]	2012-05-20
Comment[AEExperimentType]	transcription profiling by array						
Comment[GEOReleaseDate]	2012-05-19
Comment[ArrayExpressSubmissionDate]	2011-06-12
SDRF File	E-GEOD-29946.sdrf.txt