Comment[ArrayExpressAccession]	E-MTAB-3711
MAGE-TAB Version	1.1						
Investigation Title	Identification of chicken Interferon Stimulated Genes (ISGs) using Affymetrix GeneChip Chicken Genome Arrays						
Comment[Submitted Name]	Identification of chicken Interferon Stimulated Genes (ISGs) using Affymetrix GeneChip Chicken Genome Arrays
Experiment Description	Recombinant chicken IFN1 was prepared as previously reported (Laidlaw et al, 2013) and was added in chicken embryo fibroblasts to a final concentration of 1000 U/ml. Confluent cells were treated with chicken IFNa or mock treated and incubated for six hours before harvesting. The experiment was repeated in triplicate with three different batches of CEFs.						
Experimental Design	compound treatment design	replicate design	reference design				
Experimental Design Term Source REF	EFO	EFO	EFO				
Experimental Design Term Accession Number	EFO_0001755	EFO_0001776	EFO_0001775				
Experimental Factor Name	compound	dose					
Experimental Factor Type	compound	dose					
Experimental Factor Term Source REF							
Experimental Factor Term Accession Number							
Person Last Name	Giotis	Skinner					
Person First Name	Efstathios	Michael					
Person Mid Initials	S	A					
Person Email	e.giotis@imperial.ac.uk	m.skinner@imperial.ac.uk					
Person Phone	7896498097	0207 594 3938					
Person Fax	0207 594 3973 	0207 594 3973 					
Person Address	Section of Virology Faculty of Medicine, St. Mary’s Campus Norfolk Place, London, W2 1PG	Section of Virology Faculty of Medicine, St. Mary’s Campus Norfolk Place, London, W2 1PG					
Person Affiliation	Postdoctoral scientist in Virology Imperial College London	Reader in Virology Imperial College London   					
Person Roles	submitter						
Date of Experiment	2012-06-30						
Public Release Date	2015-10-01						
Protocol Name	P-MTAB-45524	P-MTAB-45525	P-MTAB-45526	P-MTAB-45527	P-MTAB-45528	P-MTAB-45529	P-MTAB-45530
Protocol Type	growth protocol	treatment protocol	nucleic acid extraction protocol	nucleic acid labeling protocol	nucleic acid hybridization to array protocol	array scanning and feature extraction protocol	normalization data transformation protocol
Protocol Term Source REF	EFO	EFO	EFO	EFO	EFO	EFO	EFO
Protocol Term Accession Number	EFO_0003789	EFO_0003969	EFO_0002944	EFO_0003813	EFO_0003815	EFO_0003814	EFO_0003816
Protocol Description	Freshly isolated CEFs were provided by the Institute of Animal Health, Compton, Berks, UK. Cells were seeded in T25 flasks (Greiner Bio One; 5.6 x 106 cells/flask) and cultured overnight in 5.5 ml 199 media (Gibco®, Invitrogen) supplemented with 8% heat-inactivated newborn calf serum (NBCS; Gibco®, Invitrogen), 10% tryptose phosphate broth (TPB; Sigma), 2% nystatin (Sigma) and 0.1% penicillin streptomycin (Gibco®, Invitrogen).	Recombinant chicken IFNα was prepared as previously reported (Laidlaw et al, 2013) and was added in culture media to a final concentration of 1000 u/ml. Confluent cells were treated with chicken IFNα or mock treated and incubated for six hours before harvesting. Cells were stored at -80°C in RNAlater (Sigma) until RNA extraction. The experiment was repeated in triplicate with three different batches of CEFs.	Total RNA was extracted from cells using an RNeasy kit (Qiagen) according to the manufacturer’s instructions. On-column DNA digestion was performed using RNase-free DNase (Qiagen) to remove contaminating genomic DNA. RNA samples were quantified using a Nanodrop Spectrophotometer (Thermo Scientific) and checked for quality using a 2100 Bioanalyzer (Agilent Technologies). All RNA samples had an RNA integrity number (RIN) ≥ 9.8.	Biotinylated fragmented RNA was prepared for each sample using standard procedures in GeneChip ® 3´ IVT Express Kit Users manual. 	Array hybridisation was performed according to the manufacturer's instructions (Affymetrix). Labeled samples were hybridized to the GeneChip Chicken Genome Arrays (Affymetrix Inc.) in a GeneChip Hybridization Oven for 16 h at 45°C and 60 rpm in an Affymetrix Hybridization Oven 645. 	After washing and staining, the arrays were scanned with the Affymetrix GeneChip Scanner 3000 7G. Gene-level expression signal estimates were derived from CEL files generated from raw data using the multi-array analysis (RMA) algorithm implemented from the Affymetrix GeneChip Command Console Software Version 3.0.1. 	Data pre-processing and filtering was done using Partek software, v.6.6 and included: RMA background correction, quantile normalization across all chips in the experiment, log2 transformation, median polish summarization.
Protocol Hardware							
Protocol Software							
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/					
Term Source Version							
Comment[AEExperimentType]	transcription profiling by array						
SDRF File	E-MTAB-3711.sdrf.txt