|
| Status |
Public on Jan 06, 2023 |
| Title |
Old Blood 1 |
| Sample type |
SRA |
| |
|
| Source name |
peripheral blood
|
| Organism |
Mus musculus |
| Characteristics |
age: old
strain: C57BL/6
tissue: peripheral blood
gender: female
|
| Growth protocol |
C57BL/6 female 4 month and 24 month mice were obtained from the National Institute of Aging (NIA). They were fed ad libitum and kept in standard housing conditions.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Blood samples were drawn from four young and four old mice. Samples from two mice within each age group were pooled, resulting in two pooled young and two pooled old samples. Subsequently, the pooled samples were diluted 1:1 with PBS+2% FBS and loaded into SepMate-15™ tubes with 4.5 mL of LymphoprepTM with a density of 1.077g/mL (StemCell Technologies). Cells were centrifuged at 1200x g for 20min at 4℃. The top layer was collected and centrifuged at 300xg for 8 min. Supernatant was removed and cells were resuspended in PBS + 2% FBS and this step was repeated. Subsequently, red blood cell lysis was performed using the Red Blood Cell Lysis Solution (Miltenyi Biotec) following the manufacturer’s protocol that includes the washing step.
Single-cell RNA-seq was performed using 10x Genomics 3' expression protocol
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
10x chromium 3' gene expression
|
| Data processing |
The Cell Ranger Single Cell Software Suite 2.1.0 was used to perform single cells demultiplexing and UMI counting (https://support.10xgenomics.com/single-cell-gene-expression/software/overview/welcome). The transcriptome reference used was mm10. Subsequently, the duplicates from young and old mice, respectively, were aggregated using Cellranger aggr. Cells that have more than 500 genes detected and less than 10% of mitochondrial reads were included in the downstream analysis using Seurat 2.3.0 . Young and old UMI counts were merged using the MergeSeurat function. Default parameters of Seurat was used in the analysis, unless otherwise stated. 1080 highly variable genes were used as an input for PCA. T-SNE projection and clustering analysis (dims=1:30 and resolution=0.4) were performed using Seurat. Markers genes for each cluster were found using the FindConservedMarkers function and the changes with age within each cell type were identified using the FindMarkers function.
Genome_build: mm10
Supplementary_files_format_and_content: Blood_UMI.txt contains UMI counts from 10x dataset and Blood_metadata.txt contains the metadata for the UMI counts
|
| |
|
| Submission date |
Sep 26, 2018 |
| Last update date |
Jan 06, 2023 |
| Contact name |
Nicola Neretti |
| Organization name |
Brown University
|
| Department |
Department of Molecular Biology, Cell Biology & Biochemistry
|
| Street address |
70 Ship St
|
| City |
Providence |
| State/province |
RI |
| ZIP/Postal code |
02903 |
| Country |
USA |
| |
|
| Platform ID |
GPL21103 |
| Series (1) |
| GSE120505 |
Single-cell RNA seq of aging peripheral blood |
|
| Relations |
| BioSample |
SAMN10132580 |
| SRA |
SRX4741345 |
