biostudies-arrayexpressMetabolomicsUnknownTranscriptomicsGenomicsProteomicsDongjuan Daitranscription profiling by arrayEscherichia coli K-12Escherichia coli K-12https://www.ebi.ac.uk/biostudies/studies/E-GEOD-24914Transcription profile of Escherichia coli cells in biofilms under static batch culture was compared to that of E. coli cells in planktonic cultures. Both E. coli biofilm and planktonic cultures were cultivated for 18 h in 10% Luria-Bertani broth at room temperature (20 degree Celsius). Biofilms were grown in static batch culture in petri dishes. Both planktonic culture and biofilms were homogenized and run through a separated protocol. Two condition experiments: E. coli biofilm vs E. coli planktonic cultures. Two biological replicates with independently grown and harvested biofilms or planktonic cultures. Each biological replicate has two technical replicates of hybridization on microarray slides. Each slide has three built-in replicates for each probe.biostudies-arrayexpressHybridization - Microarray slides were pre-hybridized with 1 mg/ml BSA in prehybridization buffer for 4 h. Slide washing, hybridization and washing were performed according to the manufacturer's protocol (Corning Epoxide Coated Slides instructions, Lowell, MA). Hybridization was at 42 °C water bath for 16 h.Sample Treatment - Planktonic cultures of E. coli were harvested and resuspended in RNAlater and kept in 4C fridge overnight. Suspended cells were removed from petri dishes and biofilms were washed three times with fresh 10% LB before being craped from the bottom surface of petri dishes. Biofilms were also re-suspeded into RNAlater and kept at 4C overnight. Both planktonic and biofilm samples were homogenized for 2 min on ice with an Omni TH homogenizer. Cells were then aliquoted into vials containing around 2x10^8 cells. Cells in each vial were then re-suspended in nuclease-free phosphate buffered saline, incubated with anti-E. coli antibody and microbeads, followed by separation on a MACS separator (Miltenyi, Auburn, CA) at 4 degree C. Sorted cells were re-suspended into RNAlater.Nucleic Acid Extraction - RNAlater was removed from cells by filtration. Total RNA was extracted from cells using a hot SDS/phenol protocol (http://www.genome.wisc.edu/pub/reprints/) with an additional 1-min bead-beating. Chemical extractions were performed in phase lock gels (Fisher Scientific, Pittsburgh, PA).Growth Protocol - Cells for inoculation were from overnight planktonic culture in 10% Luria-Bertani broth at 30 degree Celsius. Cells were washed in equal volume of fresh 10% LB broth before inoculation. Planktonic cultures were conducted in flasks with 30 ml 10% LB, inoculated with 300 ul E. coli overnight culture. Flasks were set on a shaker (250 rpm) at room temperature (20 C) for 18 h. Biofilms were cultivated in static disposable petri dishes (60 mm x 15 mm) with 5 ml 10% LB, inoculated with 50 ul E. coli overnight culture. The petri dishes were set static at room temperature (20 C) for 18 h.Scaning - Microarray slides were scanned with a Virtek ChipReader (Virtek Vision, Waterloo, ON, Canada). Spots on scanned images were recognized and pixel intensity for each spot was quantified using the TIGR software Spotfinder (v3.1.1).Labeling - RNA was reversely transcribed into cDNA using random hexamers (pd(N)6) (GE Healthcare, Piscataway, NJ) and labeled with Amersham CyDye Post-Labeling Reactive Dye (Amersham Biosciences, Piscataway, NJ) following the manufacturer's protocol provided by the Amino Allyl cDNA Labeling Kit (Ambion, Austin, TX). The two biological replicates were labeled with dye swapping.MIAME ScoreRaw DataOrganizationAssays and DataProcessed DataMAGE-TAB FilesArray DesignsDongjuan DaiChuanwu XiLutgarde RaskinData Transformation - Data was analyzed in the software Acuity 4.0 (Molecular Devices, Sunnyvale, CA). Hybridized spots for E. coli K12 having a high QC (quality control) value >0.1, good flag tags (A, B and C) in both Cy3/Cy5 channels were chosen for analysis. LOWESS normalization was performed with three iterations using a smoothing factor of 0.4. One sample t-tests were performed across replicates. P-value of 0.05 was chosen as the significant level. ID_REF = VALUE = Normalized log2 ratio of (planktonic culture/biofilm)falseEscherichia coli biofilms vs planktonic cultureTranscription profile of Escherichia coli cells in biofilms under static batch culture was compared to that of E. coli cells in planktonic cultures. Both E. coli biofilm and planktonic cultures were cultivated for 18 h in 10% Luria-Bertani broth at room temperature (20 degree Celsius). Biofilms were grown in static batch culture in petri dishes. Both planktonic culture and biofilms were homogenized and run through a separated protocol. Two condition experiments: E. coli biofilm vs E. coli planktonic cultures. Two biological replicates with independently grown and harvested biofilms or planktonic cultures. Each biological replicate has two technical replicates of hybridization on microarray slides. Each slide has three built-in replicates for each probe.2013-12-30T00:00:00Z2023-08-10T04:56:39.548Z2022-03-09T05:14:19.758ZE-GEOD-24914GSE24914EFO_0002768