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Sample GSM1134053 Query DataSets for GSM1134053
Status Public on Nov 13, 2013
Title bonemarrow-monocyte-S2-1114541 [RNA-seq]
Sample type SRA
Source name bonemarrow-monocyte
Organism Mus musculus
Characteristics tissue: bone marrow
cell type: monocyte
strain: C57BL/6
age: 6 weeks
gender: Female
Treatment protocol Tissue was dissociated, enriched by Percoll gradient, and cells were flow sorted.
Extracted molecule total RNA
Extraction protocol Total RNA was isolated from snap-frozen sorted cell pellets using TRIzol-chloroform extraction, resuspended in 10µl Ambion Nuclease-free water (Life Technologies).
For cDNA synthesis, we added 5µl of isolated RNA into the Ovation® RNA-Seq method.  500ng cDNA was fragmented using covaris shearing and processed for Illumina library construction with the Illumina paired-end LT indexing protocol as previously published (Mardis et al., 2009; Govindan et al., 2012). Each library was sequenced on the Illumina HiSeq, generating between 15-22Mbp per lane.
RNA-Seq (300-500bp size fractionation): cDNA samples were constructed into Illumina libraries according to the manufacturer’s protocol (Illumina Inc, San Diego, CA) with the following modifications: 1) DNA was fragmented using Covaris S2 DNA Sonicitor using the following condtions: Duty Cycle: 5, Intensity: 4, Cycles/Burst: 200, Time: 90sec (Covaris, Inc. Woburn, MA). Fragment sizes ranged between 100 and 500bp. 2) Illumina adapter-ligated DNA was amplified in a single 50ml PCR for five cycles. 3) Solid Phase Reversible Immobilization (SPRI) bead cleanup was used to purify the PCR and select for 300-500bp fragments. We added 0.8 volumes of the AmpureXP solution (preformulated with polyethylene glycol and sodium chloride) to size select DNA molecules greater than 300 bp. The size-fractioned cDNA was washed three times with 750 µl of 70% ethanol, the beads were dried, and the cDNA library was eluted off the beads by adding 20µl 10 mM Tris-HCl (pH 8.0). The size-fractioned libraries were assayed using the Agilent BioAnalyzer High Sensitivity DNA chips.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
Description bone marrow monocyte biological replicate 2
RNA-Seq (300-500bp size fractionation)
Data processing Basecalls performed using CASAVA version 1.7.0
Aligned with TopHat 1.3.1, Bowtie 0.12.7 and Ensembl 58 annotation (-G param)
FPKM calculated per sample with Cufflinks 1.0.3
Normalized expression calculated per sample by Alexa-Seq 1.17
Genome_build: mm9
Supplementary_files_format_and_content: Matrix_GeneExpression_v60_S1-6.txt, a tab delimited file with columns for gene ID and the normalized expression values per sample name
Supplementary_files_format_and_content: 11881_gene_fpkm.tsv, a tab delimited file with columns for gene ID and the normalized expression values per sample name
Submission date May 07, 2013
Last update date May 15, 2019
Contact name Winnie W Pong
Organization name Washington University School of Medicine
Department Department of Neurology
Lab David H Gutmann Laboratory
Street address 660 S. Euclid Avenue
City St Louis
State/province MO
ZIP/Postal code 63110
Country USA
Platform ID GPL13112
Series (2)
GSE46688 F11R is a novel monocyte prognostic biomarker for malignant glioma [Seq]
GSE46690 F11R is a novel monocyte prognostic biomarker for malignant glioma
BioSample SAMN02138463
SRA SRX275698

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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