|
| Status |
Public on Feb 15, 2021 |
| Title |
LCL_777_B958 |
| Sample type |
SRA |
| |
|
| Source name |
primary B cells (EBV-infected)
|
| Organisms |
Homo sapiens; human gammaherpesvirus 4 |
| Characteristics |
cell type: B lymphocyte
donor: 777
viral strain (transformant): B95-8
|
| Treatment protocol |
No additional treatments were applied after primary cell infection and culture
|
| Growth protocol |
Primary human B cells were isolated from whole blood by Ficoll gradient. CD19+ B cells were isolated from purified PBMCs by magnetic separation. Purified B cells were infected in bulk by incubation with viral supernatnants prepared from stimulated B95-8 Z-HT or M81 cells. Viral supernatants were incubated with primary B cells for 1 hr at 37 degrees C with 5% CO2. Cells were rinsed in 1x PBS after infection and cultured in RPMI 1640 media with 15% fetal calf serum (R15). Prior to single cell preparation, outgrown LCLs were cultured in RPMI 1640 media with 10% fetal calf serum (R10) supplemented with 2 mM L-glutamine, 100 U/mL penicillin, 100 ug/mL streptomycin, and 0.5 ug.mL cyclosporine A.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Each of the three samples was prepared separetly using the 10x Genomics Chromium controller
Standard library construction as specified by the manufacturer's protocols (v2 chemistry)
capture and single-cell indexing of total polyA mRNA
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
total polyA mRNA (single cell, host and viral)
|
| Data processing |
[base-calling] Illumina HiSeq 3000/4000
[alignment] cellranger v.2.0.0 (mkfastq); alignment was performed against the human genome (hg38) concatenated with the type 1 EBV genome (NC_007605) as an extra chromosome to capture viral transcripts
[qc filtering] Seurat v.3.1.5 (R); filtered out cells with <200 or >65000 unique RNA molecules and cells with >5% mitochondrial gene expression.
[normalization] Seurat v.3.1.5 (R)
[cell cycle marker regression] Seurat v.3.1.5 (R)
[scaling and feature selection] Seurat v.3.1.5 (R)
[PCA, dimensional reduction, and visualization] Seurat v.3.1.5 (R)
Supplementary_files_format_and_content: TSV, MTX
|
| |
|
| Submission date |
Sep 21, 2020 |
| Last update date |
Feb 15, 2021 |
| Contact name |
Elliott Daniel SoRelle |
| E-mail(s) |
elliott.sorelle@duke.edu
|
| Organization name |
Duke University
|
| Department |
Molecular Genetics & Microbiology
|
| Lab |
Micah Luftig Lab
|
| Street address |
130 Research Drive (CARL Building)
|
| City |
Durham |
| State/province |
NC |
| ZIP/Postal code |
27710 |
| Country |
USA |
| |
|
| Platform ID |
GPL23362 |
| Series (1) |
| GSE158275 |
Single-cell characterization of transcriptomic heterogeneity in lymphoblastoid cell lines |
|
| Relations |
| BioSample |
SAMN16231004 |
| SRA |
SRX9162428 |
