|
| Status |
Public on Aug 31, 2017 |
| Title |
F70_A82V_1hr |
| Sample type |
SRA |
| |
|
| Source name |
F70_MDDCs_A82V
|
| Organism |
Homo sapiens |
| Characteristics |
donor id: healthy donor #F70
cell type: monocyte-derived dendritic cells (MDDCs)
stimulation: 1 hour stimulation with A82V M-EBOV
|
| Treatment protocol |
MDDCs were seeded at 5x10^5 cells per well in 24-well plates and incubated with 100ul HIV-derived lentiviral vector pseudotyped with M-EBOV founder or A82V-containing Ebola virus glycoprotein. These challenges were performed in a total volume of 250ul culture medium. Cells were harvested at times indicated postinnoculation with virus.
|
| Growth protocol |
Peripheral blood mononuclear cells (PBMCs) were isolated by gradient centrifugation on Histopaque-1077 and CD14+ mononuclear cells were enriched via positive selection using anti-CD14 antibody MicroBead conjugates, according to the manufacturer’s protocol. CD14+ cells were then plated at a density of 1 to 2 x 106 cells/ml in RPMI-1640 supplemented with 5% heat inactivated human AB+ serum, 20 mM L-glutamine, 75 mM HEPES pH 7.2, 1 mM sodium pyruvate, and 1 x MEM non-essential amino acids. Differentiation of the CD14+ monocytes into dendritic cells (MDDCs) was promoted by addition of recombinant human GM-CSF and human IL-4; cytokines were produced from HEK293 cells stably transduced with pAIP-hGMCSF-co or pAIP-hIL4-co, respectively, as previously described (C. Reinhard et al., Retrovirology, 2014) with each cytokine supernatant added at a dilution of 1:100.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy Miniplus
Nugen Ovation FFPE RNA-seq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Because of their lower quality, the first and last base of all reads was trimmed, we then used trimmomatic-0.32 to trim by quality, removing 5’ or 3’ stretches of bases having average quality of less than 20 in a window size 10
Only reads longer than 36 bases were kept for further analysis. After quality trimming reads were aligned to the human ribosomal RNA using Bowtie2 v 2.2.3 with parameters -p 2 -N 1 --no-unal
All reads mapped to rRNA were discarded from further analysis. To estimate gene and isoform expression in Transcripts per Million (TPM) units and raw counts (expected_count), we used RSEM v1.2.28
Genome_build: hg19
Supplementary_files_format_and_content: tsv, read counts and tpm normalized counts
|
| |
|
| Submission date |
Jul 26, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Alper Kucukural |
| E-mail(s) |
alper.kucukural@umassmed.edu
|
| Phone |
7743124493
|
| Organization name |
UMass Medical School
|
| Department |
Program in Molecular Medicine
|
| Lab |
Biocore
|
| Street address |
364 Plantation Street
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE84865 |
Ebola virus glycoprotein variant with increased infectivity for human cells dominated the 2013-2016 outbreak |
|
| Relations |
| BioSample |
SAMN05441364 |
| SRA |
SRX1978350 |
