 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jan 30, 2024 |
| Title |
Mid thoracic spinal cord, 077_A (spatial) |
| Sample type |
SRA |
| |
|
| Source name |
Mid thoracic spinal cord
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
tissue: spinal cord
condition: Mixture of uninjured, 7 days post SCI, 2 months post SCI
genotype: wild type
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For snRNA-seq experiment, single-nucleus dissociation of the mouse lumbar spinal cord was performed according to our established procedures. Following euthanasia by isoflurane inhalation and cervical dislocation, the spinal cord injury site was immediately dissected and frozen on dry ice. Spinal cords were doused in 500 µL sucrose buffer (0.32 M sucrose, 10 mM HEPES [pH 8.0], 5 mM CaCl2, 3 mM Mg acetate, 0.1 mM EDTA, 1 mM DTT) and 0.1% Triton X-100 with the Kontes Dounce Tissue Grinder. 2 mL of sucrose buffer was then added and filtered through a 40 µm cell strainer. The lysate was centrifuged at 3200 g for 10 min at 4°C. The supernatant was then decanted, and 3 mL of sucrose buffer was added to the pellet for 1 min. We homogenized the pellet using an Ultra-Turrax and 12.5 mL of density buffer (1 M sucrose, 10 mM HEPES [pH 8.0], 3 mM Mg acetate, 1 mM DTT) was added below the nuclei layer. The tube was centrifuged at 3200 g at 4°C and supernatant poured off. Nuclei on the bottom half of the tube wall were collected with 100 µL PBS with 0.04% BSA and 0.2 U/µL RNase inhibitor. Finally, nuclei were resuspended through a 30 µm strainer, and adjusted to 1000 nuclei/µL. snRNA-seq library preparation was carried out using the 10x Genomics Chromium Multiome ATAC + Gene Expression Kit Version 3. The nuclei suspension was added to the Chromium RT mix to achieve loading numbers of 10,000 nuclei. For downstream cDNA synthesis (13 PCR cycles), library preparation and sequencing, the manufacturer’s instructions were followed. For spatial transcriptomics, we carried out two separate experiments to study the cytoarchitecture of the lesion microenvironment after SCI. First, we prepared sections from uninjured mice, 7 days and 2 months after crush SCI. For each experimental condition, we prepared sections from the lesion epicenter of three independent biological replicates. Second, to prepare our four-dimensional spatiotemporal atlas, we collected sections throughout the entire spinal cord of mice from each of the three experimental conditions. The spinal cord injury sites of mice were embedded in OCT and cryosections were generated at 10 µm at −20 °C. For the four-dimensional atlas, every fifth section was collected throughout the entire dorsoventral axis of each spinal cord. Sections were immediately placed on chilled Visium Tissue Optimization Slides or Visium Spatial Gene Expression Slides.
We used 10x Genomics Chromium Single Cell Gene Expression Kit Version 3.1 kit to profile the snRNA sequencing and Visium Spatial Gene Expression to profile the spatial transcriptomics.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
10x v3
|
| Data processing |
For the snRNA-seq, reads were aligned using Cellranger v 4.0.0 and the final data matrix is generated as per Cellranger pipeline. For the spatial transcriptomics, reads were aligned with SpaceRanger v 1.0.0 and the final data matrix is generated as per Spaceranger pipeline
Data were analyzed using Seurat v3
Assembly: mm10-1.2.0 (GRCm38.101)
Supplementary files format and content: tab-delimited text files include UMI values for each cell with tab delimeters
Library strategy: Spatial Transcriptomics
|
| |
|
| Submission date |
Jun 12, 2023 |
| Last update date |
Jan 30, 2024 |
| Contact name |
Alan Yue Yang Teo |
| E-mail(s) |
yueyang.teo@epfl.ch
|
| Organization name |
EPFL
|
| Department |
SV
|
| Lab |
UPCOURTINE
|
| Street address |
Chem. des Mines 9, Campus Biotech
|
| City |
Geneve |
| ZIP/Postal code |
1202 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL17021 |
| Series (1) |
| GSE234774 |
The Tabulae Paralytica: Multimodal single-cell and spatial atlases of spinal cord injury |
|
| Relations |
| BioSample |
SAMN35720253 |
| SRA |
SRX20664522 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7474508_077_A_scalefactors_json.json.gz |
170 b |
(ftp)(http) |
JSON |
| GSM7474508_077_A_tissue_hires_image.png.gz |
5.1 Mb |
(ftp)(http) |
PNG |
| GSM7474508_077_A_tissue_lowres_image.png.gz |
428.7 Kb |
(ftp)(http) |
PNG |
| GSM7474508_077_A_tissue_positions_list.csv.gz |
54.4 Kb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
