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Sample GSM8269454 Query DataSets for GSM8269454
Status Public on May 22, 2024
Title control diet, replicate 2
Sample type SRA
Source name brain
Organism Mus musculus
Characteristics tissue: brain
cell type: sorted endothelial cells
treatment: control diet
treatment duration: 1 month
strain: C57BL6
Treatment protocol Mice were kept on control or PLX5622 (1200 ppm) diet for one month
Extracted molecule total RNA
Extraction protocol Mice were anesthetized with ketamine/xylazine and decapitated. Brains were removed, meninges were peeled off, and cerebellum and olfactory bulbs were discarded. A series of enzymatic and mechanical steps was used to achieve a single cell suspension of the remaining brain tissue. FACS was used to enrich for CD31+ endothelial cells, which were sorted for single-cell sequencing. Brains from three mice were used across the two samples.
Libraries were constructed with the 10X Genomics Chromium Next GEM Single Cell 3’ kit v3.1 according to manufacturer's instructions
Library strategy RNA-Seq
Library source transcriptomic single cell
Library selection cDNA
Instrument model Illumina NovaSeq 6000
Data processing Gene counts were obtained by aligning reads to the mm10 genome (refdata-gex-mm10-2020-A) using CellRanger software (v.6.0.1 – 10X Genomics). Using Seurat (v.4.0), quality control was performed using the following: 1) outliers with a high ratio of mitochondrial RNAs (>5%, <200 features) relative to endogenous RNAs and homotypic doublets (>5000 features) were removed in Seurat; 2) after scTransform normalization and integration, doublets and multiplets were removed using DoubletFinder; 3) cells were manually inspected using known cell-type specific marker genes, cells expressing more than one cell-type specific marker were removed.
In Seurat (v.4.0), gene counts were first normalized using scTransform, then Integration function was used to align data with default settings. Genes were projected into principal component (PC) space using the principal component analysis function RunPCA. The first 30 dimensions were used as inputs into Seurat’s FindNeighbors and FindClusters functions. Then, RunUMAP function with default settings was used to calculate 2-dimensional UMAP coordinates and search for distinct cell populations.
Distinct cell types were determined using known cell-type specific markers. Differential gene expression of genes comparing PLX5622 to control samples was performed with the FindMarkers function, specifically utilizing the Wilcoxon Rank Sum test.
Assembly: mm39
Supplementary files format and content: barcodes, features, and matrix for each sample
Submission date May 15, 2024
Last update date May 22, 2024
Contact name Caterina Profaci
Organization name University of California, San Diego
Street address 9500 Gilman Drive
City La Jolla
ZIP/Postal code 92093
Country USA
Platform ID GPL24247
Series (1)
GSE267591 Microglia are not necessary for maintenance of blood-brain barrier properties in health, but PLX5622 alters brain endothelial cholesterol metabolism (Brain Endo SC Seq)
BioSample SAMN41411759
SRA SRX24569754

Supplementary file Size Download File type/resource
GSM8269454_C2_barcodes.tsv.gz 2.6 Mb (ftp)(http) TSV
GSM8269454_C2_features.tsv.gz 254.1 Kb (ftp)(http) TSV
GSM8269454_C2_matrix.mtx.gz 58.9 Mb (ftp)(http) MTX
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Raw data are available in SRA

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