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Sample GSM3130812 Query DataSets for GSM3130812
Status Public on Jun 21, 2018
Title LPS_IC_DMSO_REP2_004
Sample type SRA
 
Source name Bone marrow
Organism Mus musculus
Characteristics strain: C57BL/6
tissue: Bone marrow
age: 8-10 weeks
Treatment protocol BMDMs were primed overnight with recombinant murine IFNg (10 ng/mL) (BioLegend, USA) and washed twice by media and warm PBS, respectively. Salmonella LPS (100 ng/mL) (Sigma-Aldrich, USA), rabbit IgG against ovalbumin (OVA) (GeneTex, USA) and IgG-opsonized OVA (immune complex) were added to activate macrophages as indicated. Gamma secretase inhibitor (GSI), DAPT (25 µM) (Calbiochem, USA) and vehicle control DMSO (0.01%) (Sigma-Aldrich, USA) were added to macrophages.
Growth protocol To generate bone marrow derived macrophages (BMDMs),bone marrow cells were plated in non-tissue culture treated plates (Hycon, Thailand) and cultured at 37°C, 5% CO2 in 8 mL of media containing DMEM supplemented with 10% Fetal Bovine Serum, HEPES, sodium pyruvate , streptomycin/penicillin G (all from LONZA, USA or Hyclone, UK), 5% (v/v) horse serum (Thermo Scientific, USA) and 20% (v/v) L929-conditioned media. Three mL of fresh DMEM media supplemented with 20% L929-conditioned media and 5% horse serum was added to the culture at day 4. Cells were harvested at the day 7 and used for all the experiments.
Extracted molecule total RNA
Extraction protocol Total RNA samples were extracted from LPS/IC-activated BMDMs in the presence of DMSO or GSI for 1 hr using an RNeasy Mini Kit (Qiagen, USA). RNA samples were assessed for quality using an Agilent 2100 Bioanalyzer (Agilent, USA) and quantified using a Qubit 3.0 fluorometer (Life Technologies, USA).
RNA libraries were prepared for sequencing using standard Illumina protocols
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NextSeq 500
 
Data processing bcl2fastq2 Conversion Software v2.20 used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and then mapped to mm9 whole genome using STAR version 2.5.0a with parameters --runMode alignReads --runThreadN 12 --readFilesCommand zcat --outStd Log --outSAMunmapped within --outSAMtype BM SortedByCordinate limitGenomeGenerateRAM 40,000,000 --limitIObufferSize 40,000,000 --outWigType wiggle --chimSegmentMin 21
bam files were merge using picard and remove duplicate or PCR product contamination
Raw count from each samples were generated using HT-Seq program with parameters --HTSeq.GenomicArrayOfSets("auto", stranded = True)
Genome_build: mm10
Supplementary_files_format_and_content: tab-limited files with raw counts value
 
Submission date May 03, 2018
Last update date Jun 21, 2018
Contact name Tanapat Palaga
E-mail(s) tanapat.p@chula.ac.th
Phone 6622185070
Organization name Chulalongkorn University
Department Microbiology
Lab 2015
Street address Payathai Road, Pathumwan
City Bangkok
ZIP/Postal code 10330
Country Thailand
 
Platform ID GPL19057
Series (1)
GSE114020 Effect of Notch Signaling Inhibitor on Gene Expression in Lipopolysaccharide-stimulated Macrophages in the Presence of Immune Complex
Relations
BioSample SAMN09060761
SRA SRX4037335

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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