Comment[ArrayExpressAccession] E-GEOD-48527 MAGE-TAB Version 1.1 Public Release Date 2014-08-14 Investigation Title DNA methylation and gene expression analysis of an Epithelial-to-mesenchymal transition model Comment[Submitted Name] DNA methylation and gene expression analysis of an Epithelial-to-mesenchymal transition model Experiment Description This SuperSeries is composed of the SubSeries listed below. Refer to individual Series Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Gómez Person First Name Antonio Person Email agomezm@idibell.cat Person Affiliation IDIBELL Person Address PEBC, IDIBELL, Hospital Duran i Reynals Av. Gran Via s/n km, 2.7, L'Hospitalet de Llobregat, Barcelona, Spain Person Roles submitter Protocol Name P-GSE48527-1 P-GSE48527-8 P-GSE48527-9 P-GSE48527-5 P-GSE48527-2 P-GSE48527-4 P-GSE48527-7 P-GSE48527-6 P-GSE48527-3 P-GSE48527-10 Protocol Description Expression Console v1.2 (Affymetrix) ID_REF = VALUE = RMA normalizationn applied ATLAS Biolabs GmbH ATLAS Biolabs GmbH treatment with TGFβ (R&D systems, Minneapolis, MN) at 5 ng/ml during 30 days, subculturing the cells every 3-4 days Treatment with TGFbeta at 5 ng/ml (RD Systems) DNA extracted by Phenol:Chloroform:Isoamylalcohol (Sigma) BS-seq library constructing, 10ug genomic DNA was fragmented using a Covaris sonication system (Covaris S2). Following fragmentation, libraries were constructed using the Illumina Paired-End protocol consisting of end repair, base addition and methylated-adaptor ligation. Ligated DNA was bisulfite converted using the EZ DNA Methylation-Gold kit (ZYMO). Different insert size were excised from the same lane of a 2% TAE agarose gel. Normally, three bands are excised corresponding to DNA insert sizes of 80-100 bp, 100-120 bp to 120-150bp. Products were purified by using QIAquick Gel Extraction kit (Qiagen) and amplified by PCR. PCR was carried out in a final reaction volume of 50ul consisting of 20ul purified DNA, 4ul 2.5 mM dNTP, 5 ul 10X buffer, 0.5 ul JumpStart Taq DNA polymerase, 2ul 10uM PCR primers and 37.5 ul ÎœltraPure TM Water and the following thermal cycling program: 94C 30 s, 10 cycles of 94C 30 s, 60C 30 s, 72C 30 s then prolong with 1 min at 72C. PCR products were sequenced with an Illumina genome analyzer. The reads generated by Illumina sequencing were aligned to the reference genome using SOAPaligner2. The alignment and methylation estimation was performed as described before (Li, Y. et al. PLoS Biol 8, e1000533 (2010)). 100 bases Pair-end reads. TRIZOL and Isopropanol precipitation was used for RNA extraction DMEM 10% FBS, standard CO2/O2/temperature conditions Cells were cultured in DMEM at 10% FBS, 5% CO2. Control cells were maintained in culture during 30 days, in parallel with the TGFbeta-treated cells. ATLAS Biolabs GmbH Protocol Type normalization data transformation protocol labelling protocol hybridization protocol sample treatment protocol sample treatment protocol nucleic acid library construction protocol nucleic acid library construction protocol growth protocol growth protocol array scanning protocol Experimental Factor Name CELL PHENOTYPE COMPOUND Experimental Factor Type cell phenotype compound Comment[SecondaryAccession] GSE48527 Comment[GEOReleaseDate] 2014-08-14 Comment[ArrayExpressSubmissionDate] 2013-07-03 Comment[GEOLastUpdateDate] 2014-08-15 Comment[AEExperimentType] transcription profiling by array Comment[AEExperimentType] methylation profiling by high throughput sequencing SDRF File E-GEOD-48527.hyb.sdrf.txt E-GEOD-48527.seq.sdrf.txt