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| Status |
Public on Jun 07, 2019 |
| Title |
naïve |
| Sample type |
SRA |
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| Source name |
PBMCs
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| Organism |
Homo sapiens |
| Characteristics |
infection: control
time (post-infection): 0 hr
cell type: PBMCs
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| Treatment protocol |
Salmonella strain used in this study was derived from the wild-type strain SL1344 containing GFP (pFPV25.1; Addgene). Cultures of Salmonella were grown in Luria-Bertani (LB) medium at 37°c for 16 hours and used for PBMCs infection at MOI 25 for the exposed cells, and PBS was added to the naive samples. After 30 min of internalization, the cells were washed and suspended with media containing 50 ug/ml gentamicin to eliminate Salmonella that were not internalized. The cells were incubated for 4 hours at 37°c in 5% CO2 in non-treated cell culture plates.
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| Growth protocol |
Venous blood was drawn from the cubital vein of volunteers and PBMCS were isolated as described in Li et al. Cell 2016. The cells were counted and frozen until used. A day before experiment, the cells where defrosted, suspended in medium (RPMI 1640 with L- Glutamine supplemented with 10% heat inactivated fetal bovine serum and 1mM sodium pyruvate) and plated on untreated plates. A day after, the cells were collected from the dish. To avoid cell lost, the dish was washed with medium and the remaining cells were added to the collected cells. The cells were then manually counted with trypan blue.
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| Extracted molecule |
total RNA |
| Extraction protocol |
4 hours after infection, the cells were washed with PBS, counted with trypan blue, suspended with 0.04% BSA in PBS and directly used for single-cell sequencing by the Chromium Single Cell 3’ Reagent version 2 kit and Chromium Controller (10X Genomics, CA, USA) as previously described at Zheng, G. X. Y. et al Nature Communications 2017.
Libraries were prepared by the Chromium Single Cell 3’ Reagent version 2 kit and Chromium Controller (10X Genomics, CA, USA) as previously described at Zheng, G. X. Y. et al Nature communications 2017.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Reads: R1 contains the cell barcode (16 nt) and UMI (10 nt). R2 contains the cDNA sequence. I1 contains the sample index.
The Cell Ranger 2.1.1 Single-Cell Software Suite (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger) was used to perform sample demultiplexing, alignment to the genome (GRCh38), barcode assignment for each cell and gene counting by unique molecular identifier (UMI) counts.
Overall we sequenced 7000 cells; 3515 cells from the naïve sample, with ~80,500 mean reads per cell, ~1800 median UMI count per cell and ~800 median genes per cells. For the exposed sample we sequenced 3485 cells with ~76,000 mean reads per cell, ~1800 median UMI count per cell and ~830 median genes per cells.
Only genes with at least one UMI count detected in at least one cell were used
Data was normalized to a library size factor. Factors were calculated by dividing total UMI counts in each cell to the median of the total UMI counts across all cells
Data was transformed to log10 scale (log10(UMI count+1))
Genome_build: GRCh38
Supplementary_files_format_and_content: [*cells.txt ] the UMI count of each gene in each cell
[data_matrix.txt] the normalized UMI counts of the genes in the cells from both samples (naive and exposed; 7000 cells).
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| Submission date |
Nov 01, 2018 |
| Last update date |
Jun 07, 2019 |
| Contact name |
Noa Bossel Ben-Moshe |
| Organization name |
Weizmann
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| Street address |
Hertzl Street
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| City |
Rehovot |
| ZIP/Postal code |
76100 |
| Country |
Israel |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE122083 |
Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [scRNA-seq ind.1] |
| GSE122084 |
Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells |
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| Relations |
| BioSample |
SAMN10359534 |
| SRA |
SRX4967250 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3454528_naive_cells.txt.gz |
4.2 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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