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| Status |
Public on Jun 07, 2019 |
| Title |
TLR10_3_t4_rep3 |
| Sample type |
SRA |
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| Source name |
PBMCs
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| Organism |
Homo sapiens |
| Characteristics |
genotype: TLR10
individual (ind name): TLR10#3
infection: Salmonella enterica serovar Typhimurium
time (post-infection): 4 hr
cell type: PBMCs
replicate: 3
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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 the time indicated in each experiment 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. 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 and normalized so the same amount of cells was used for each individual (1-5*10^5 cells per replicate, depends on the amount of live cells in the least concentrated sample).
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| Extracted molecule |
total RNA |
| Extraction protocol |
At the indicated time points after infection the cells were washed with PBS, then suspended with RLT buffer (from RNeasy Mini Kit, Qiagen) + 1% β-mercaptoethanol and kept in -80°c until further extraction. RNA was then extracted with RNeasy Mini Kit (Qiagen).
RNA-seq libraries were prepared according to Cel-seq libraries protocol (Hashimshony, T. et al Genome Biology 2016) with a minor modification: the starting material was purified bulk RNA.
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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 |
The cel-seq pipeline (https://github.com/yanailab/CEL-Seq-pipeline) was used for sample demultiplexing, alignment to the genome (GRCh38), and gene counting.
Data was normalized to a library size factor. Factors were calculated by dividing the total number of reads from each sample to the median of the total number of reads across all samples.
Data was transformed to log2 scale, and minimal expression threshold was set to 2.
The three replicates of each sample were averaged, except for 3 samples we excluded due to low coverage (<100K exonic reads), and therefore for 3 samples (TLR10#1 t=4h, TLR10#2 t=0 and WT#3 t=0), only 2 replicates were averaged.
Genome_build: GRCh38
Supplementary_files_format_and_content: [*rep*txt] the processed normalized gene count table of each replicate
[data_matrix.txt] the average expression levels of each sample (from 3 replicates)
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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) |
| GSE122058 |
Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [WT/TLR10 bulk RNA-seq] |
| GSE122084 |
Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells |
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| Relations |
| BioSample |
SAMN10358923 |
| SRA |
SRX4964989 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3453913_TLR10_3_t4_rep3.txt.gz |
75.8 Kb |
(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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