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| Status |
Public on Apr 01, 2024 |
| Title |
50% WT GFP+ BMDMs and 50% N4bp1 KO BMDMs unstimulated |
| Sample type |
SRA |
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| Source name |
Bone Marrow
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| Organism |
Mus musculus |
| Characteristics |
background: C57BL/6
tissue: Bone Marrow
genotype: N4bp1 knockout
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| Treatment protocol |
Treatment with TLR7 for 8h
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| Extracted molecule |
total RNA |
| Extraction protocol |
Bone marrow cells from femurs and tibia of wild-type C57BL/6N mice were cultured in Roswell Park Memorial Institute (RPMI) medium supplemented with 10% heat-inactivated fetal bovine serum, 2 mM glutamine, 50 μM 2-mercaptoethanol (Gibco), 100 U/ml penicillin, and 100 μg/ml streptomycin (Gibco) and 50 ng/ml of recombinant M-CSF (R&D Systems) on non-tissue culture treated 15 cm plates. On day 5 of culture, cells were harvested and 5 million cells were re-plated onto non-tissue culture treated 10 cm plates in fresh media (same as that described above). After 2 additional days of culture, BMDMs from individual mice were harvested by scraping.
scRNAseq library constructions were carried out using the 10X Chromium Controller and the Chromium Next GEM Single Cell 3’ Reagent Kits (V3.1) in accordance with the manufacture’s protocol. Prior to loading into the chromium controller, cells were washed once with PBS and resuspended in PBS at 700-1200 cells/ul for a target cell recovery of 7000 cells.
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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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| Description |
SAM24397624
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| Data processing |
10X immune profiling single-cell RNA sequencing data were processed with our in-house pipeline. Briefly, the expression library were first demultiplexed followed by alignment to the custom mouse reference genome (GRCm38 + GFP sequence) to measure the expression of GFP using Cell Ranger (version 6.0.1). A gene by cell barcode matrix was generated and the downstream analysis was performed using Seurat.
Seurat (version 4.0.4) was used for all downstream analysis. Cells with detected genes between 500 and 5000, and with less than 5% mitochondrial mapped reads were kept for downstream analysis. After filtering, we detected 36,174 cells across the whole dataset. After normalizing the data, we performed linear dimensionality reduction using PCA. We ranked the principal components (PCs) by the amount of variance explained by each and found that after the first 20 PCs the explained variance plateaus. To visualize the data in two-dimensional space, Uniform Manifold Approximation and Projection (UMAP) was performed using the first 20 PCs. We then used the UMAP dimensions to determine the nearest neighbor of each cell and perform clustering by setting the resolution parameter to 0.15.
Assembly: mm10
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| Submission date |
Nov 29, 2022 |
| Last update date |
Apr 01, 2024 |
| Contact name |
Rohit Reja |
| E-mail(s) |
rejar@gene.com
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| Phone |
+1-650-467-7615
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| Organization name |
Genentech
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| Street address |
1 DNA Way
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| City |
South San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94080 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE218957 |
Regulation of IkB kinase family crosstalk by an N4BP1-caspase-8 axis [scRNA-seq] |
| GSE218958 |
Regulation of IkB kinase family crosstalk by an N4BP1-caspase-8 axis |
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| Relations |
| BioSample |
SAMN31924512 |
| SRA |
SRX18410650 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6760839_SAM24397624_matrix.mtx.gz |
120.4 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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