{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["Dongjuan Dai"],"study_type":["transcription profiling by array"],"organism":["Escherichia coli K-12"],"species":["Escherichia coli K-12"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-GEOD-24912"],"description":["Transcription profile of Escherichia coli cells in mono-species pure biofilms was compared to that of E. coli cells in E. coli-Stenotrophomonas maltophilia dual-species biofilms. E. coli cells were separated from dual-species biofilms before total RNA extraction to eliminate possible cross hybridization from S. maltophilia transcripts. The separation method was developed by combining the use of reagent RNAlater and immuno-magnetic separation. Pure E. coli biofilms were processed with the same separation protocol before RNA extraction. Two condition experiments: E. coli mono-species biofilm vs E. coli in mixed-species biofilm. Two biological replicates with independently grown and harvested biofilms. Each biological replicate has two or three technical replicates of hybridization on microarray slides. Each slide has three built-in replicates for each probe."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Sample Treatment - Suspended cells were then removed and biofilms were washed 3 times with 1 ml 10% LB broth. Biofilms from each petri dish were then scraped into a vial containing 1 ml 10% LB. Scraped biofilms in each vial were concentrated from 1 ml into 50 ul by centrifugation. 500 ul RNAlater was added and mixed well. Samples in RNAlater were kept in 4C fridge overnight. Each vial was homogenized with OMNI TH homogenizer for 2 min on ice and further aliquoted into small vials. Cells in each vial were then re-suspended in nuclease-free phosphate buffered saline, incubated with anti-E. coli antibody (ViroStat) and microbeads (Miltenyi), followed by separation on a MACS separator (Miltenyi, Auburn, CA) at 4 degree C. Sorted cells were re-suspended into RNAlater until RNA extractions.","Hybridization - 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.","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 and were inoculated into 5 ml 10% LB in a disposable petri dish (60 mm x 15 mm). 50 ul E. coli culture was inoculated for mono-species pure culture. 50 ul E. coli and 10 ul S. maltophilia were mixed and inoculated for mixed-species culture. Petri dishes were set static at room temperature (20 C) for 18 h for biofilm growth.","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).","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."],"figure_sub":["MIAME Score","Raw Data","Organization","Assays and Data","Processed Data","MAGE-TAB Files","Array Designs"],"pubmed_authors":["Dongjuan Dai","Chuanwu Xi","Lutgarde Raskin"],"data_protocol":["Data 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 (mixed-species biofilm/pure mono-species biofilm)"],"additional_accession":[]},"is_claimable":false,"name":"Escherichia coli biofilm: mono-species culture vs. mixed-species culture","description":"Transcription profile of Escherichia coli cells in mono-species pure biofilms was compared to that of E. coli cells in E. coli-Stenotrophomonas maltophilia dual-species biofilms. E. coli cells were separated from dual-species biofilms before total RNA extraction to eliminate possible cross hybridization from S. maltophilia transcripts. The separation method was developed by combining the use of reagent RNAlater and immuno-magnetic separation. Pure E. coli biofilms were processed with the same separation protocol before RNA extraction. Two condition experiments: E. coli mono-species biofilm vs E. coli in mixed-species biofilm. Two biological replicates with independently grown and harvested biofilms. Each biological replicate has two or three technical replicates of hybridization on microarray slides. Each slide has three built-in replicates for each probe.","dates":{"release":"2013-12-30T00:00:00Z","modification":"2023-08-11T15:07:24.073Z","creation":"2022-01-28T19:31:33.291Z"},"accession":"E-GEOD-24912","cross_references":{"GEO":["GSE24912"],"EFO":["EFO_0002768"]}}