Comment[ArrayExpressAccession]	E-MTAB-3712
MAGE-TAB Version	1.1
Investigation Title	Identification of chicken Interferon Stimulated Genes (ISGs) using Affymetrix Chicken Gene 1.0 ST Arrays
Comment[Submitted Name]	Identification of chicken Interferon Stimulated Genes (ISGs) using Affymetrix Chicken Gene 1.0 ST 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 IFNα 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	reference design	replicate design
Experimental Design Term Source REF	EFO	EFO	EFO
Experimental Design Term Accession Number	EFO_0001755	EFO_0001775	EFO_0001776
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	07896498097	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	2014-07-10
Public Release Date	2015-10-01
Protocol Name	P-MTAB-45531	P-MTAB-45532	P-MTAB-45533	P-MTAB-45534	P-MTAB-45535	P-MTAB-45536	P-MTAB-45537
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.	Labeling of samples for array analysis was performed using the GeneChip WT Terminal labeling assay (Affymetrix) with 100 ng input RNA.	Array hybridisation was performed according to the manufacturer's instructions (Affymetrix). Labeled samples were hybridized to the Affymetrix Chicken Gene 1.0 ST Arrays in a GeneChip Hybridization Oven for 16 h at 45°C and 60 rpm. 	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 normalisation across all chips in the experiment, log2 transformation, median polish summarisation.
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-3712.sdrf.txt