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Sample GSM2236774 Query DataSets for GSM2236774
Status Public on May 15, 2020
Title poly(A) mRNA MDSC_p50ko_IFNg R2
Sample type SRA
Source name Monocytic Myeloid Derived Suppressor Cells (M-MDSC) replicate 2
Organism Mus musculus
Characteristics strain: C56Bl6/J
genotype: p50ko
cell type: M-MDSC (28 post injection)
treatment: Interferon-treated
Treatment protocol 5 million M-MDSC were plated in RPMI1640 containing 10% FBS and left untreated or stimulated with 200U/ml interferon gamma for 4 hours.
Growth protocol CD11b+Ly6C+ Monocytic Myeloid Derived Suppressor Cells (M-MDSC) were obtained by serial selection using magnetic separation from tumor-bearing C56Bl6/J mice, 28 days after injection of murine fibrosarcoma (MN/MCA1). In details, myeloid suppressor populations were first enriched by serial negative selections with CD19 and CD11c microbeads, and then positively selected with Ly6G microbeads. Remaining cells were positively selected with CD11b+ microbeads. The purity of the cell populations evaluated by flow cytometry exceeded 90%.
Extracted molecule polyA RNA
Extraction protocol The poly-A containing mRNA molecules were purified using poly-T oligo-attached magnetic beads. Following purification, the mRNA was fragmented into small fragments using divalent cations and elevated temperature. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This was followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. cDNA fragments were end-repaired and a single ‘A’ base was added before adapter ligation. The products were then purified and enriched by PCR to create the final cDNA library.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NextSeq 500
Data processing Reads were adapter-trimmed according to the Illumina pipeline (Bcl2fastq)
Reads were mapped to the mouse mm9 genome and transcripts using TopHat 1.3.1 (PMID: 19289445), allowing up to two mismatches and with a mean distance between pairs (-r) of 150bp
Transcripts abundance were quantified using HTSeq (PMID: 25260700)
Differentially expressed genes were called using edgeR (PMID: 19910308)
Genome_build: mm9 (NCBI Build 37), transcripts from Mus Musculus UCSC iGenome
Supplementary_files_format_and_content: Counts per million (CPM) mapped reads obtained using edgeR
Submission date Jul 15, 2016
Last update date May 15, 2020
Contact name Viviana Piccolo
Organization name European Institute of Oncology (IEO)
Department Department of Experimental Oncology
Lab Gioacchino Natoli Lab
Street address Via Adamello 16
City Milano
State/province Milano
ZIP/Postal code 20139
Country Italy
Platform ID GPL19057
Series (1)
GSE84468 Tumor-derived prostaglandin E2 promotes p50 NF-κB-dependent differentiation of monocytic MDSC.
BioSample SAMN05408028
SRA SRX1951576

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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