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Sample GSM1695697 Query DataSets for GSM1695697
Status Public on May 27, 2015
Title RNA-seq TH-ProB cultured 1 day on OP9DL1, rep2
Sample type SRA
Source name Ebf1+/- Pax5+/- ProB, 1 day culture on OP9DL1 stromal cells
Organism Mus musculus
Characteristics strain/background: C57BL/6J
genotype/variation: Ebf1+/- Pax5+/-
cell type: ProB
treatment: 1 day culture on OP9DL1 stromal cells
Treatment protocol See above.
Growth protocol ProB-cell isolation / culture protocol: For sorting of Pro B cells, CD16/CD32 (FC) -blocked (93, eBioscience) cells were stained with antibodies against lineage markers CD11b/Mac1 (M1/70), Gr1 (RB6-8C5), TER119 (Ter119), CD3 (17A2, BD Pharmingen), CD11c (N418) and NK1.1 (PK136) and further stained with CD19 (ID3), CD45R/B220 (RA3-6B2), CD43 (S7), IgM (RMM-1) and IgD (11-26. eBioscience, San Diego, CA). Cell sorting was performed on a BD FACSAriaTM (BD Biosciences, San Jose, California) using propidium iodide (PI, Invitrogen, Paisly, UK) as viability marker. For expansion, Pro-B cells were cultured on OP9 stromal cells supplemented with 10ng/mL KIT ligand, 10ng/mL Fms-like tyrosine kinase 3 ligand (FLT3L), and 10ng/mL Interleukin-7 in Optimem supplemented with 10% FCS, Beta-mercaptoethanol, HEPES and Penstrep.
Extracted molecule total RNA
Extraction protocol RNA-Seq: RNA was extracted with the Qiagen RNeasy Micro Kit.
RNA-seq 1 day on OP9DL1 and OP9: Libraries were constructed using Nugen's Ovation Ultralow Library systems. Detailed protocol is available on their website. In short, input DNA were fragmented using sonication, end repaired, followed by ligation of adapters, amplification and finally bead purification. The samples were run on NextSeq 500.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NextSeq 500
Data processing For RNA-Seq samples, data analysis was performed with Arraystar (DNA STAR). The reads were aligned to mouse reference genome (mm9, NCBI 37) and RPKM normalized. Statistical analysis in Arraystar was performed with StudentĀ“s t-test with correction for multiple testing (Benjamini Hochberg). A p-value > 0.05 was considered as statistically significant.
Genome_build: MGSCv37 (mm9)
Supplementary_files_format_and_content: Processed data files include BED files. All genomic coordinates are relative to the mm9 mouse assembly.
Submission date May 26, 2015
Last update date May 15, 2019
Contact name Rajesh Somasundaram
Phone 0046708890787
Organization name Linkoping University
Department Microbiology and Molecular Medicine
Lab Lab 1, Floor- 13 Dept of Clinical and Experimental Medicine (IKE)
Street address Dept of Clinical and Experimental Medicine (IKE)
City Linkoping
ZIP/Postal code SE-58185
Country Sweden
Platform ID GPL19057
Series (1)
GSE69227 Combined heterozygous loss of Ebf1 and Pax5 allows for T-lineage conversion of B-cell progenitors
BioSample SAMN03734560
SRA SRX1038505

Supplementary file Size Download File type/resource
GSM1695697_ProB_TH_OP9DL1-2.bed.gz 57.9 Mb (ftp)(http) BED
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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