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Sample GSM1382099 Query DataSets for GSM1382099
Status Public on Jun 04, 2014
Title mEpiSC_RNAseq_rep2
Sample type SRA
 
Source name Epiblast stem cells
Organism Mus musculus
Characteristics cell line: EpiSC7
Growth protocol mESCs were grown in Knockout DMEM (Invitrogen) supplemented with 20% Knockout Serum Replacement (KSR) (Invitrogen), 10^3 units/ml recombinant murine Leukemia Inhibitory Factor (LIF) (ESGRO, Millipore), 2mM L- alanyl-L-glutamine dipeptide (Glutamax; Invitrogen), 1x non-essential amino acids (Invitrogen), and 0.1mM 2-mercaptoethanol (Sigma). mESCs were passaged every third day as a single cell suspension using 0.25% trypsin/EDTA and seeded at 3.0x10^4 cells/cm^2 for routine culture. mEpiSCs were cultured in Knockout DMEM supplemented with 20% KSR, 10ng/ml FGF2 (R&D Systems), 2mM Glutamax, 1x non-essential amino acids, and 0.1mM 2-mercaptoethanol and passaged every third day using 1.5 mg/ml collagenase type IV (Invitrogen) and trituration into small clumps of ~10-100 cells. Irradiated mouse embryonic fibroblasts (MEFs) served as a feeder layer for all pluripotent cell types and were maintained with DMEM supplemented with 10% fetal bovine serum, 2 mM glutamax, 1x non-essential amino acids, and 0.1 mM 2- mercaptoethanol. All cells were grown on Nunclon Δ-treated dishes or multiwell plates (Fisher Scientific) coated for 2 hours at 37°C with 0.1% (w/v) porcine gelatin (Sigma).
Extracted molecule polyA RNA
Extraction protocol Cells were separated from MEF feeder layer and RNA was extracted from ~2x10^6 cells with TRIzol (Invitrogen), separated using Phase Lock Gel tubes (5 Prime), and purified using the RNAeasy Plus kit (Qiagen) according to the manufacturer’s protocol.
PolyA+ RNA was prepared for sequencing using the Illumina TruSeq RNA Sample Preparation Kit according to the manufacturer's protocol.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Data processing Reads were aligned to mm9 genome build (retrieved from http://cufflinks.cbcb.umd.edu/igenomes.html), using Tophat v2.0.6 (Trapnell et al., 2009) with default settings.
FPKM values for RefSeq genes were calculated using Cufflinks v2.0.2 (Trapnell et al., 2010) provided with the GTF file via the -G (known genes only) option.
Genome_build: mm9
Supplementary_files_format_and_content: tsv files - tab-delimited files containing FPKM values for RefSeq genes in each sample generated using Cufflinks v2.0.2 (Trapnell et al., 2010). A detailed description of the file format can be found at http://cufflinks.cbcb.umd.edu/manual.html under the subheading "Cufflinks Output"
 
Submission date May 07, 2014
Last update date May 15, 2019
Contact name Paul J. Tesar
E-mail(s) paul.tesar@case.edu
Organization name Case Western Reserve University School of Medicine
Department Genetics and Genome Sciences
Street address 10900 Euclid Ave
City Clev
State/province OH
ZIP/Postal code 44106
Country USA
 
Platform ID GPL13112
Series (2)
GSE57403 Epigenomic comparison of distinct pluripotent stem cell states reveals a new class of enhancers with roles throughout mammalian development (RNA-seq)
GSE57409 Epigenomic comparison of distinct pluripotent stem cell states reveals a new class of enhancers with roles throughout mammalian development
Relations
Reanalyzed by GSE80797
BioSample SAMN02767287
SRA SRX535196

Supplementary file Size Download File type/resource
GSM1382099_mEpiSC_RNAseq_rep2_genes.tsv.gz 782.4 Kb (ftp)(http) TSV
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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