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Sample GSM4774469 Query DataSets for GSM4774469
Status Public on Oct 13, 2020
Title 5_RNAseq_mESC_serum_rep2
Sample type SRA
 
Source name E14 mouse embryonic stem cells
Organism Mus musculus
Characteristics modification: 3xFlag knock-in to endogeous Kdm2b C-terminus
genotype: wild-type
strain: 129P2/OlaHsd
Growth protocol The wild-type E14 mESCs with 3xFlag knock-in to the endogenous Kdm2b gene were maintained on the 0.1% gelatin coated plates in "2i" medium that contained 50% Neurobasal medium (Life Technologies) and 50% DMEM/F12 medium (Life Technologies) supplemented with 1x N2-supplement (Life Technologies), 1x B27 supplement (Life Technologies), 7.5% bovine serum albumin (Life Technologies), 1x GlutaMAX (Life Technologies), 1x beta-mercaptoethanol (Life Technologies), 1mM PD0325901, 3mM CHIR99021, 1000 units/ml leukemia inhibitory factor (ESGRO, EMD Millipore), and 100U/ml penicillin/streptomycin (Life Technologies). The PD culture conditoin is the same as 2i culture condtion except for it containng only 1mM PD0325901. The serum-containing mESC culture condition included DMEM medium (Life Technologies) supplemented with 100U/ml penicillin/streptomycin (Life Technologies), 15% fetal bovine serum (Sigma), 1x nonessential amino acid, 1x sodium pyruvate (Life Technologies), 1x GlutaMAX (Life Technologies), 1x beta-mercaptoethanol (Life Technologies) and 1000 units/ml leukemia inhibitory factor (ESGRO, EMD Millipore).
Extracted molecule total RNA
Extraction protocol RNA was extracted and purified from cells using QI shredder (Qiagen) and RNeasy (Qiagen) spin columns.
Total RNA (1 µg) was used to generate RNA-seq library using NEBNext Ultra Directional RNA library Prep Kit for Illumina (New England BioLabs, Inc) according to the manufacturer’s instructions. Adapter-ligated cDNA was amplified by PCR and followed by size selection using agarose gel electrophoresis. The DNA was purified using Qiaquick gel extraction kit (Qiagen) and quantified both with an Agilent Bioanalyzer and Invitrogen Qubit. The libraries were diluted to a working concentration of 10nM prior to sequencing. Sequencing on an Illumina HiSeq4000 instrument was carried out by the Genomics Core Facility at Michigan State University.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 4000
 
Description serum medium
1_cuffdiffout_mESC_serum_2i_PD_gene_exp.diff
Data processing For the RNA-Seq data analysis, all sequencing reads were mapped to NCBI build 37 (mm9) of the mouse genome using Tophat2.
The mapped reads were normalized to reads as Reads Per Kilobase of transcript per Million mapped reads (RPKM).
The differential gene expression was calculated by Cuffdiff program and the statistic cutoff for identification of differential gene expression is q < 0.05 and 1.5-fold RPKM change between samples.
Genome_build: mm9
 
Submission date Sep 09, 2020
Last update date Oct 13, 2020
Contact name Jin He
E-mail(s) hejin1@msu.edu
Phone 5173530613
Organization name Michigan State Univeristy
Street address 603 Wilson Road, Biochem Bldg. Rm. 410
City East Lansing
State/province Michigan
ZIP/Postal code 48824
Country USA
 
Platform ID GPL21103
Series (2)
GSE157747 Cell signaling coordinates global Polycomb Repressive Complex 2 recruitment and developmental gene expression in murine embryonic stem cells (RNA-Seq)
GSE157749 Cell signaling coordinates global Polycomb Repressive Complex 2 recruitment and developmental gene expression in murine embryonic stem cells
Relations
BioSample SAMN16090757
SRA SRX9101809

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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