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Sample GSM2450583 Query DataSets for GSM2450583
Status Public on Apr 25, 2023
Title 96h-rep1
Sample type SRA
Source name E14
Organism Mus musculus
Characteristics strain: 129/Ola
treatment: 96 hours retinoic acid + N2B27
facs: Not sorted
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.
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).
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.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 3000
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.
Submission date Jan 09, 2017
Last update date Apr 26, 2023
Contact name Stefan Semrau
Organization name Leiden University
Department Physics
Lab Quantitative Single Cell Biology
Street address Einsteinweg 55
City Leiden
ZIP/Postal code 2333CC
Country Netherlands
Platform ID GPL21493
Series (1)
GSE93301 Integration of a multi-omics stem cell differentiation dataset using a dynamical model
BioSample SAMN06212450
SRA SRX2480218

Supplementary file Size Download File type/resource
GSM2450583_96h-rep1_TAGCTT-6.genes.results.txt.gz 940.7 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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