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Sample GSM1025307 Query DataSets for GSM1025307
Status Public on Feb 06, 2013
Title macrophages_treated_with_LPS+PGE_1hr_rep_3
Sample type RNA
 
Source name primary bone marrow derived mouse macrophages, treated with LPS and PGE, harvested after 1 hour of treatment
Organism Mus musculus
Characteristics strain: C57Bl/6 wild type mouse
cell type: primary macrophages
tissue: bone marrow
Treatment protocol BMDMs were either left untreated or stimulated with 100 ng/ml LPS (#L6529; Sigma) or a combination of 100 ng/ml LPS and 10 microM prostaglandin E for 1 hour
Growth protocol To obtain primary bone marrow derived mouse macrophages (BMDMs), bone marrow cells were maintained on bacterial grade plates for 1 week in DMEM supplemented with 10% heat inactivated FBS (Biosera), 2 mM L-glutamine, 100 units/ml penicillin G, 100mg/ml streptomycin, 0.25 mg/ml amphotericin (Invitrogen) and 5 ng/ml mCSF (R&D systems). Adherent cells were then re-plated on tissue culture grade plates in fresh media and used 24 hours after re-plating. RAW264.7 cells were cultured in DMEM supplemented with 10% heat inactivated FBS, 2 mM L-glutamine, 100 units/ml penicillin G and 100mg/ml streptomycin.
Extracted molecule total RNA
Extraction protocol RNA extraction was carried out by homogenisation of BMDMs in Trizol followed by phenol chloroform extraction.
Label biotin
Label protocol 250 ng of total RNA was used from each sample. The total RNAs were processed with Ambion WT Expression kit (P/N 4411973) and Affymetrix Genechip® WT Terminal Labelling and Controls Kit (P/N 901524), according to manufactures protocols. RNA concentration was checked with Nanodrop ND-2000 and Agilent Bioanalyzer electrophoresis station was used for quality control.
 
Hybridization protocol Samples were hybridized to Affymetrix Mouse Gene 1.1 ST array plate using manufacturer's protocols for using the GeneTitan® Hybridization, Wash and Stain Kit for WT Array Plates (P/N 901622)
Scan protocol GeneTitan® Instrument was used to hybridize, wash, stain and scan the arrays. Affymetrix GeneChip® Command Console® (AGCC) 3.1 was used to control GeneTitan® hybridization process and in summarizing probe cell intensity data (.CEL file generation). The hybridization quality was checked with AGCC and Expression Console 1.1.
Data processing The data analysis was carried out using Affymetrix Power Tools (APT), R (version 2.13.1) – Bioconductor and Partek GS 6.5 (version 6.11.0321) software. For quality control and probesets annotations, the latest annotations files (Release 32, dated 23-06-2011) downloaded from Affymetrix were used. Using APT, The probe level expression data were summarised to transcript cluster level, filtered to include DABG (detected above background) and normalised using two different normalisation methods – RMA and Plier-GCBA (GC background-adjusted). The dataset quality was analysed from spike in probesets, principal component analysis, hierarchical clustering analysis and by generating various quality control plots such as mean raw signal intensity plots, mean absolute deviation (MAD) of the residuals plots and probe cell intensity plots. The processed dataset submitted here (matrix table) shows quality control passed, DABG (detected above background) filtered data summarised to transcript cluster level and normalised using RMA normalisation method using Affymetrix Power Tools.
 
Submission date Oct 25, 2012
Last update date Feb 06, 2013
Contact name Manikhandan A V Mudaliar
E-mail(s) innovatebio@gmail.com
Organization name University of Glasgow
Department College of Medical, Veterinary and Life Sciences
Lab Institute of Infection, Immunity and Inflammation
Street address Room B528, Sir Graeme Davies Building, 120 University Place
City Glasgow
State/province Scotland
ZIP/Postal code G12 8TA
Country United Kingdom
 
Platform ID GPL11533
Series (1)
GSE41833 Expression profiles of primary bone marrow derived mouse macrophages – untreated and treated with LPS or LPS+PGE

Data table header descriptions
ID_REF
VALUE RMA normalised, log2 transformed and DABG (detected above background) filtered transcript cluster level signal

Data table
ID_REF VALUE
10338001 11.29206
10338002 4.863
10338003 9.5504
10338004 6.41496
10338006 2.46907
10338007 2.93522
10338009 6.76379
10338011 4.344
10338013 1.91762
10338017 12.53379
10338018 5.10838
10338019 4.04776
10338025 6.45401
10338026 12.81177
10338027 5.53853
10338028 3.7214
10338029 9.19291
10338032 2.0189
10338033 5.16594
10338034 3.14336

Total number of rows: 35309

Table truncated, full table size 585 Kbytes.




Supplementary file Size Download File type/resource
GSM1025307_1h_PGE_LPS3_C05.CEL.gz 4.2 Mb (ftp)(http) CEL
Processed data included within Sample table

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