|
| Status |
Public on Jun 03, 2022 |
| Title |
Patient 9247 10x VDJ Replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
PBMC
|
| Organism |
Homo sapiens |
| Characteristics |
individual: Patient 9247
age: 31
Sex: Male
hiv-1 infected: Yes
aviraemic: Yes
art: Yes
tissue: PBMC
cell type: CD4+ T cells
library type: VDJ RNA
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
PBMCs were obtained from HIV-1 infected, aviraemic individuals under ART by leukapheresis and stored on liquid nitrogen. CD4+ CD45RA- cells were isolated by magnetic-activated cell sorting. Cells were subsequently incubated with Fc blocking reagent, and stained with a combination of anti-CD3 and/or anti-CD4, anti-TRBC1, and one or a combination of various anti-TRBV antibodies. Cells were sorted for CD3 and/or CD4, a specific TRBC domain and/or a specific TRBV domain or a subset of various TRBV domains.
Library was performed according to the manufacter’s instructions (single cell 5’ and V(D)J Enrichment v2 protocol, 10x Genomics). Briefly, GCs were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
| |
|
| Description |
5' VDJ library: read1 file contains cell barcode and UMI; read2 file contains transcript
|
| Data processing |
The demultiplexing, barcoded processing, TCR assembly and gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: hg38
Supplementary files format and content: 10x Genomics output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz, VDJ_filtered_contig_annotations.csv
Library strategy: VDJ-seq
|
| |
|
| Submission date |
May 24, 2022 |
| Last update date |
Jun 03, 2022 |
| Contact name |
Thiago Y Oliveira |
| E-mail(s) |
toliveira@rockefeller.edu
|
| Organization name |
The Rockefeller University
|
| Department |
Immunology, Virology and Microbiology
|
| Lab |
Laboratory of Molecular Immunology
|
| Street address |
1230 YORK AVE
|
| City |
NEW YORK |
| State/province |
New York |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
|
| Platform ID |
GPL21697 |
| Series (1) |
| GSE204756 |
Distinct gene expression by expanded clones of quiescent memory CD4+ T cells harboring intact latent HIV-1 proviruses |
|
| Relations |
| BioSample |
SAMN28647790 |
| SRA |
SRX15446298 |
