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GEO help: Mouse over screen elements for information. |
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Status |
Public on Apr 25, 2023 |
Title |
72h |
Sample type |
SRA |
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Source name |
E14
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Organism |
Mus musculus |
Characteristics |
strain: 129/Ola treatment: 72 hours retinoic acid + N2B27 facs: Not sorted
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Treatment protocol |
E14 cells were treated with Retinoic acid to induce differentiation. Prior to differentiation cells were grown in 2i medium for at least 2 passages. Cells were seeded at 2.5 × 105 per 10 cm dish and grown over night (12 h). Cells were then washed twice with PBS and basal N2B27 medium (2i medium without the inhibitors, LIF and the additional insulin) supplemented with 0.25 μM all-trans retinoic acid (RA, Sigma-Aldrich). Spent medium was exchanged with fresh medium after 48 h.
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Growth protocol |
E14 mouse embryonic stem cells were grown in modified 2i medium: DMEM/F12 (Life technologies) supplemented with 0.5x N2 supplement, 0.5x B27 supplement, 4mM L- glutamine (Gibco), 20 μg/ml human insulin (Sigma-Aldrich), 1x 100U/ml penicillin/streptomycin (Gibco), 1x MEM Non-Essential Amino Acids (Gibco), 7 μl 2-Mercaptoethanol (Sigma-Aldrich), 1 μM MEK inhibitor (PD0325901,Stemgent), 3 μM GSK3 inhibitor (CHIR99021, Stemgent), 1000 U/ml mouse LIF (ESGRO). Cells were passaged every other day with Accutase (Life technologies) and replated on gelatin coated tissue culture plates (Cellstar, Greiner bio-one).
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Extracted molecule |
total RNA |
Extraction protocol |
Cells were dissociated with accutase. In addition, the ECT and XEN samples were sorted with fluorescence activated cell sorting (CD24+/PDGFRA- and CD24-/PDGFRA+ resp.). RNA was extracted from the samples (RNeasy, Qiagen) and the purified RNA was stored at -80C until RNA-sequencing was performed. The libraries for were prepared under the standard conditions using Illumina’s TruSeq stranded mRNA sample preparation kit. The libraries were sequenced using Illumina HiSeq 3000; 40 basepair long, stranded single-end reads were sequenced at an average read depth of 40 million reads per sample.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 3000 |
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Data processing |
Sequenced reads were trimmed for adaptor sequence and low-quality sequence. Trimmed reads were aligned to the transcriptome of mm10 with RSEM v1.2.28 and bowtie2 v2.2.6 with the options ‘--sampling-for-bam’ and ‘--fragment-length-mean 40’. To obtain unique counts for the CDS and 3’UTR of genes the BAM files were overlapped with the CDS and 3’UTR regions of mm10 using in the GenomicFeatures package in R Genome_build: mm10 Supplementary_files_format_and_content: The *.genes.results files are plain-text output files from RSEM and contain TPM and expected_count. CDS_3UTR_counts.txt contains a matrix of unique read counts of the CDS and 3'UTR in the samples.
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Submission date |
Jan 09, 2017 |
Last update date |
Apr 26, 2023 |
Contact name |
Stefan Semrau |
Organization name |
Leiden University
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Department |
Physics
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Lab |
Quantitative Single Cell Biology
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Street address |
Einsteinweg 55
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City |
Leiden |
ZIP/Postal code |
2333CC |
Country |
Netherlands |
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Platform ID |
GPL21493 |
Series (1) |
GSE93301 |
Integration of a multi-omics stem cell differentiation dataset using a dynamical model |
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Relations |
BioSample |
SAMN06212451 |
SRA |
SRX2480217 |
Supplementary file |
Size |
Download |
File type/resource |
GSM2450582_72h_GATCAG-6.genes.results.txt.gz |
952.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
Processed data are available on Series record |
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