|
| Status |
Public on Jan 08, 2014 |
| Title |
Subject6 WNV |
| Sample type |
SRA |
| |
|
| Source name |
macrophages, WNV
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: monocyte-derived macrophages
treatment: West Nile virus (WNV)
donor identifier: 6
|
| Treatment protocol |
Virulent WNV (CT-2741) was passaged once in Vero cells (ATCC CCL-81) and viral PFU were quantified by plaque assays. Cells were infected with WNV (multiplicity of infection, MOI = 1) for 24 h.
|
| Growth protocol |
Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll-Hypaque (GE Healthcare, NJ) and suspended in RPMI-1640 containing L-Glutamine supplemented with 20% (v/v) human serum (Lonza, MD), 100 U/ml penicillin and 100 µg/ml streptomycin (Invitrogen, CA). Cells were plated at 2x10e7 cells/10-cm tissue culture dish. Non-adherent cells were washed away after 2 h, and adherent monocytes were incubated 6-8 days to mature into macrophages.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
mRNA was fragmented into 150-300bp fragments by incubation in RNA Fragmentation Reagent (Ambion, TX) at 70°C for 5 minutes. The fragmented mRNA was purified away from the fragmentation buffer by Agencourt RNAClean beads (Beckman Coulter, MA) following the manufacturer's instructions. The purified, fragmented mRNA was converted into double-stranded cDNA using the SuperScript Double-Stranded cDNA Synthesis Kit (Invitrogen, CA) by priming with random hexamers.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Data processing |
Using the fastq files, all sequencing reads were mapped back to human genome (hg19) using TopHat (v1.1.4).
Cufflinks (v0.9.3) was used to assemble the mapped reads against the ENSEMBL (release 57) gene structure annotation, and estimate expression levels for each transcript.
Genome_build: hg19
Supplementary_files_format_and_content: Tab-delimited text files include RPKM values for each Sample.
|
| |
|
| Submission date |
Sep 09, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Ruth R Montgomery |
| E-mail(s) |
[email protected]
|
| Organization name |
Yale University School of Medicine
|
| Department |
Internal Medicine
|
| Street address |
300 Cedar St.
|
| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06520 |
| Country |
USA |
| |
|
| Platform ID |
GPL9115 |
| Series (1) |
| GSE40718 |
Identification of genes critical for resistance to infection by West Nile virus using RNA-Seq analysis |
|
| Relations |
| SRA |
SRX185651 |
| BioSample |
SAMN01163682 |
