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Sample GSM2469582

Query DataSets for GSM2469582

Status

Public on Jan 31, 2017

Title

s04-suscep-infect

Sample type

SRA

Source name

monocyte-derived dendritic cells

Organism

Homo sapiens

Characteristics

cell type: monocyte-derived dendritic cells
individual: s04
status: susceptible
treatment: infected
gender: male

Treatment protocol

We infected the DCs with Mycobacterium tuberculosis (MTB) H37Rv at a multiplicity of infection of 1-to-1 for 18 hours.

Growth protocol

We isolated mononuclear cells from the whole blood samples using Ficoll-Paque centrifugation, extracted monocytes via CD14 positive selection, and differentiated the monocytes into dendritic cells (DCs) by culturing them for 5 days in RPMI 1640 (Invitrogen) supplemented with 10% heat-inactivated FCS (Dutscher), L-glutamine (Invitrogen), GM-CSF (20 ng/mL; Immunotools), and IL-4 (20 ng/mL; Immunotools).

Extracted molecule

polyA RNA

Extraction protocol

We extracted RNA using the Qiagen miRNeasy Kit.
We prepared sequencing libraries using the Illumina TruSeq Kit.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Data processing

We mapped reads to human genome hg38 (GRCh38) using Subread (v1.5.0-p2) and discarded non-uniquely mapping reads (flag -u).
We downloaded the exon coordinates of 19,800 Ensembl protein-coding genes (Ensembl 83, Dec 2015, GRCh38.p5) using the R(3.2.1)/Bioconductor(3.2) package biomaRt (2.26.1) and assigned mapped reads to these genes using featureCounts (v1.5.0-p2).
We filtered genes based on their expression level by removing all genes with a transformed median log2 counts per million (cpm) of less than zero. This step resulted in a set of 11,336 genes for downstream analysis.
Genome_build: hg38
Supplementary_files_format_and_content: Supplementary_Data_S1.tds (tab-delimited) contains information on the 50 samples. Most variables describe the batch processing steps. “id” is a unique identifier for each sample, “individual” is the individual identifier (“s” = susceptible, “r” = resistant), “status” is the susceptibility status, “treatment” is if the sample was infected or non-infected, “infection” is the date of the infection experiment (12 total), “arrival” is the identifier for the arrival batch (4 total), “extraction” is the batch for RNA extraction (5 total), “master_mix” is the batch for library preparation (3 total), “rin” is the RNA Integrity Number from the Agilent Bioanalyzer, and “outlier” is a Boolean variable indicating if the sample was identified as an outlier and removed from the analysis.
Supplementary_files_format_and_content: Supplementary_Data_S2.tds (tab-delimited) contains the gene expression counts for the 11,336 genes after filtering lowly expressed genes for all 50 samples. Each row is a gene labeled with its Ensembl gene ID. Each column is a sample. Each sample is labeled according to the pattern “x##-status-treatment”, where x is “r” for resistant or “s” for susceptible, ## is the ID number, status is “resist” for resistant or “suscep” for susceptible, and treatment is “noninf” for non-infected or “infect” for infected.

Submission date

Jan 26, 2017

Last update date

May 15, 2019

Contact name

John D Blischak

Organization name

University of Chicago

Department

Human Genetics

Lab

Gilad