characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2015-09-22T17:00:53Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T18:06:45Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2570567
characteristics.characteristic: External Id
characteristics.text: EMBL EBI
characteristics.characteristic: INSDC center name
characteristics.text: 2015-09-22T17:00:53Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T18:06:45Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS471008
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-2600:a2i_90
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: G4
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0002081
characteristics.characteristic: cell line
characteristics.text: mESC
characteristics.characteristic: cell type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Cells were grown in one of 3 different media: Serum-containing media: Knockout DMEM (Gibco), 1X penicillin-streptomycin-glutamine (Gibco), 1X non-essential amino acids (Gibco), 100 U/ml recombinant human leukemia inhibitory factor (Millipore), 15% fetal bovine serum (HyClone), 0.1mM _-mercaptoethanol (Sigma). Standard 2i media: N2B27 basal media (NDiff 227, StemCells), 100 U/ml recombinant human leukemia inhibitory factor (Millipore), 1 _M PD0325901 (Stemgent), 3 _M CHIR99021 (Stemgent). Alternative 2i media: N2B27 basal media (NDiff 227, StemCells), 100 U/ml recombinant human leukemia inhibitory factor (Millipore), 1 _M CGP77675 (Sigma), 3 _M CHIR99021 (Stemgent). (SMARTer) reverse transcription and cDNA preamplification were performed in the 10-17 microns Fluidigm C1 Single-Cell Auto Prep IFC using the SMARTer PCR cDNA Synthesis kit (Clontech) and the Advantage 2 PCR kit (RNAEasy) RNA was extracted usin RNAEasy kit, Qiagen (Nextera XT) For each culture condition, 4000 cells were loaded onto a 10-17 micron Fluidigm C1 Single-Cell Auto Prep IFC and cell capture was performed according to the manufacturer's instructions. Nextera libraries were according to “Using the C1TM Single-Cell Auto Prep System to Generate mRNA from Single Cells and Libraries for Sequencing”. (TruSeq) Bulk mRNA sequencing libraries were prepared and sequenced at the Wellcome Trust Sanger Institute sample preparation pipeline using Illumina's TruSeq RNA Sample Preparation v2 kit. RNA was extracted from 1-2mln cells using Qiagen RNA Purification Kit on QiaCube robot. Quality of RNA sample was checked using gel electrophoresis. For library preparation poly-A RNA was purified from total RNA using oligo-dT magnetic pull-down, subsequently mRNA was fragmented using metal-ion catalyzed hydrolysis. cDNA was synthesized using random priming, end repair was performed to obtain blunt ends and A-tailing was done to enable subsequent ligation of Illumina paired-end sequencing adapters. Samples were multiplexed at this stage. Resulting library was amplified using 10 cycles of PCR using Kapa Hifi polymerase instead of polymerase provided by Illumina in TruSeq kit
characteristics.characteristic: description
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_10090
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.characteristic: scientific_name
characteristics.text: fresh specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000971
characteristics.characteristic: specimen with known storage state
characteristics.text: a2i_90
characteristics.characteristic: title