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| Status |
Public on Jun 04, 2014 |
| Title |
mEpiSC_RNAseq_rep4 |
| Sample type |
SRA |
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| Source name |
Epiblast stem cells
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| Organism |
Mus musculus |
| Characteristics |
cell line: EpiSC9
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| 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).
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| 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.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| 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"
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| Submission date |
May 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Paul J. Tesar |
| E-mail(s) |
paul.tesar@case.edu
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| Organization name |
Case Western Reserve University School of Medicine
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| Department |
Genetics and Genome Sciences
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| Street address |
10900 Euclid Ave
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| City |
Clev |
| State/province |
OH |
| ZIP/Postal code |
44106 |
| Country |
USA |
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| 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 |
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| Relations |
| Reanalyzed by |
GSE80797 |
| BioSample |
SAMN02767297 |
| SRA |
SRX535198 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1382101_mEpiSC_RNAseq_rep4_genes.tsv.gz |
784.4 Kb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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