|
| Status |
Public on Mar 24, 2023 |
| Title |
NEU_WT_NI_01 |
| Sample type |
SRA |
| |
|
| Source name |
Neutrophils
|
| Organism |
Mus musculus |
| Characteristics |
infection: Not infected
genotype: Wild-type M. marinum M-strain
strain: C57Bl/6JRj
|
| Treatment protocol |
C57Bl/6JRj female 8 weeks old mice were infected via tail vein injections with 5 x 10^7 bacteria per mouse, in a total volume of 200 µL.
|
| Growth protocol |
Bacteria (WT and dRD1 M. marinum) were inoculated from frozen stocks and grown at 30°C in 7H9 culture medium until late log phase.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Two independent experiments were performed. For each experiment 14 to 15 tails (i.e. infected tissue) per group were collected, pooled and digested at two weeks post infection. Cell surface markers were stained with antibodies. Live infected neutrophils (CD45+Ly6G+CD11b+Wasabi+) or live uninfected neutrophils (CD45+Ly6G+CD11b+Wasabi-) were sorted by FACS in PBS supplemented with 1% BSA using a FACS ARIA Fusion (BD Biosciences).
Single cell RNA-sequencing libraries were prepared according to the manufacturer’s instructions using Chromium Single Cell 3′ Library & Gel Bead Kit v3 (10x Genomics, PN-1000092) and Chromium Chip B Single Cell Kit (PN-1000074) with the Chromium Controller & Next GEM Accessory Kit (10x Genomics, PN-120223). Library quality control and quantification were performed using a KAPA Library Quantification Kit for Illumina Platforms (Kapa Biosystems, KK4873) and the 2100 Bioanalyzer equipped with a High Sensitivity DNA kit (Agilent, 5067-4626).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
single cell RNA sequencing analysis of infected or bystander (noninfected) neutrophils during M. marinum infection
|
| Data processing |
Demultipexing and alignment of reads with CellRanger v2.1.1 for replicates 01 and v3.1.0 for replicates 02. Data is paired and pooled on 4 lanes.
Cells with very low counts of reads and genes (2-300) dependent on individual sample distribution, and mitonchondiral gene load > 10 % were discarded.
Contaminating cell types were removed from the merged replicates before merging all samples all done w. Seurat v3.
Genome_build: mm10 (replicates 01) and mm10-2.1.0 (replicates 02)
Supplementary_files_format_and_content: tsv.gz & mtx.gz
|
| |
|
| Submission date |
Apr 14, 2021 |
| Last update date |
Mar 24, 2023 |
| Contact name |
Line Wulff |
| Organization name |
Technical University of Denmark
|
| Department |
Health Technology
|
| Lab |
Agace
|
| Street address |
Kemitorvet, Building 204
|
| City |
Lyngby |
| ZIP/Postal code |
2800 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE172072 |
The role of ESX-1 on infected and uninfected neutrophils in Mycobacterium marinum infection |
|
| Relations |
| BioSample |
SAMN18741738 |
| SRA |
SRX10601621 |
