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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Oct 20, 2021 |
| Title |
HAS3_KO_LESION |
| Sample type |
SRA |
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| Source name |
Atherosclerosis plaques
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| Organism |
Mus musculus |
| Characteristics |
cell type: Mixture of cell types (SMC, Epithelial, Macrophage, others)
strain: C57BL/6
diet: Western Diet
molecule subtype: single-cell total RNA
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| Treatment protocol |
Animals were maintened in a Western Diet for 18 weeks
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| Extracted molecule |
total RNA |
| Extraction protocol |
Individual atherosclerotic plaques from the brachiocephalic artery were removed from underlying media with forceps and deposited into FACS buffer (1% BSA in PBS) plus 1 ug/mL Actinomycin D,(Gibco, #11805017). Samples were then minced and digested with 1mL Liberase (Roche, #355374) plus 1ug/mL ActinomycinD for 1 hour at 37 degrees, then resuspended in PBS plus 0.04% UltraPure non-Acetylated BSA (ThermoFisher #AM2616) and filtered through 30um filters on ice. Samples were then either counted and submitted directly for scRNAseq or sorted by flow cytometry. Samples for flow were stained with Sytox Blue viability dye and sorted with a BD FACSAria Fusion cell sorter with gating on live, single cells that were eYFP+. Cells were collected into LoBind tubes containing 0.04% non-acetylated BSA.
2,000 cells in each group were targeted in Chromium 10X genomics libraries, which after barcoding, were pooled and sequenced on the Illumina NextSeqTM, 150 cycle high-output.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina MiSeq |
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| Data processing |
Basecalls performed using CASAVA
Reads were aligned against mouse genome mm10 using the CellRanger software (10x Genomics).
Gene-barcode matrices were analyzed in R using Seurat v342. Cells were filtered for 200-5000 reads per UMI, 10% or less mitochondrial and less than 5% hemoglobin gene content.
Significant principal components of variation (PCs) were calculated using JackStraw test with 10000 repetitions, and clusters were calculated with 19 PCs
Differential expression analysis was done using MAST
Genome_build: mm10
Supplementary_files_format_and_content: Cell-gene-barcode matrices; gz file containing barcodes, genes and matrix.mtx (10x output filtered folder)
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| Submission date |
Aug 25, 2021 |
| Last update date |
Oct 20, 2021 |
| Contact name |
Gary K Owens |
| Organization name |
University of Virginia
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| Street address |
415 Lane Road MR5 Building Room 1322
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| City |
Charlottesville |
| State/province |
VA |
| ZIP/Postal code |
22908 |
| Country |
USA |
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| Platform ID |
GPL16417 |
| Series (1) |
| GSE182795 |
scRNA-seq of atherosclerotic plaques of Apoe-/- mouse (with or without specific SMC KO of Has3) |
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| Relations |
| BioSample |
SAMN20981894 |
| SRA |
SRX11911851 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5537130_KO_filtered_feature_bc_matrix.tar.gz |
19.3 Mb |
(ftp)(http) |
TAR |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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