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| Status |
Public on Jan 11, 2024 |
| Title |
IL-10R/CerS2 DKO CD45.2 |
| Sample type |
SRA |
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| Source name |
colon
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| Organism |
Mus musculus |
| Characteristics |
tissue: colon
cell type: colon macrophage
genotype: IL-10R/CerS2 DKO donor into CD45.1 recipient
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| Extracted molecule |
total RNA |
| Extraction protocol |
single cells were isolated with collagenase D from colons of chimeric mice engrafted with either IL-10R KO or IL-10R/CerS2 DKO Cd45.2 donors. Single cells were sorted with a BD Aria FACS sorter for CD11b/C11c/CD64+ cells. Sorted cells were processed with 10x Chromium Controller
scRNAseq libraries were generated using the Chromium Next GEM Single Cell 3’ v3.1 Library & Gel Bead Kit (10X Genomics) according to the manufacturer’s protocol. Briefly, emulsions were generated using the 10X Chromium Controller (10X Genomics) for a targeted recover 10,000 cells. Barcoded cDNAs were isolated from the emulsion and amplified by PCR (11 cycles). The cDNAs underwent fragmentation, end repair, and A-tailing before addition of sample indexes by PCR (16 cycles).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Cellranger v6.0.1 was used to align scRNA-seq samples to the mouse genome (mm10).
Data analysis was performed using Seurat v4.3.0 R package, including cell-type identification and comparative analyses between conditions.
In the step of quality control, poor-quality cells with the number of expressed genes < 300 or > 5000 were filtered out. We also excluded cells if their mitochondrial gene percentages were over 10%. After combining cells from all samples, we first normalized the raw count matrix and then defined 2000 top variable genes. We then applied principal component analysis (PCA) for dimensionality reduction and retained 30 leading principal components for cell clustering.
After identifying cluster-specific genes, we annotated cell types based on canonical marker genes (Lyz2 for macrophage) and focused on macrophage populations in the downstream analyses.
To test the differential expression of inflammatory genes between IL-10R KO and Cers2/IL-10R DKOs, we first applied ALRA to impute the gene expression profiles and then used the non-parametric Wilcoxon rank sum test to get their p-values.
Assembly: mm10
Supplementary files format and content: "barcodes.tsv.gz" contains cell barcodes, "features.tsv.gz" contains metadata for genes, "matrix.mtx.gz" contains digital expression matrix
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| Submission date |
Jan 04, 2024 |
| Last update date |
Jan 11, 2024 |
| Contact name |
Rihao Qu |
| Organization name |
Yale University
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| Street address |
300 George Street
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| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06511 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (2) |
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| Relations |
| BioSample |
SAMN39262853 |
| SRA |
SRX23102474 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8003018_DKO_barcodes.tsv.gz |
4.3 Kb |
(ftp)(http) |
TSV |
| GSM8003018_DKO_features.tsv.gz |
272.8 Kb |
(ftp)(http) |
TSV |
| GSM8003018_DKO_matrix.mtx.gz |
3.8 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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