 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Sep 14, 2021 |
| Title |
mTBI2 |
| Sample type |
SRA |
| |
|
| Source name |
micro-dissections of the nRT and the adjacent relay thalamic nuclei
|
| Organism |
Mus musculus |
| Characteristics |
tissue: reticular thalamic nucleus
genotype: WT
strain: C57BL/6
treatment: mild traumatic brain injury
|
| Treatment protocol |
controlled cortical impact; mild traumaic brain injury was perfomed with a CCI device using the following parameters: 3mm tip diamtere, 15 angle, depth 0.8mm from the dura, velocity 3m/s and dwell time 100ms. Sham animals recviered identical surgical prep and cranotomy, but the injury was not delivered.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Nuclei were isolated from the nRT/thalamus. Tissue samples were homogenized with 10 strokes of the loose "A" pestle and 15 strokes of the tight "B" sized pstle. The lysate was passed through a 40uM FlowMi strainer and nuclei were peleted at 500 RCF at 4C. Pelleted nuclei were resuspended in the homogenization buffer, purified using an iodixanol gradient and immidiately used for snRNA-seq.
SnRNA-seq libraries were processed using the Chromium Next GEM Single Cell 3'v3 library kit with dual indexing according ot the manufactuere's specifications. The chromium was loaded with 9,9000 nuclei.One GEM rxn was used for each sample. Replicate 1 and 2 nuclei were processed on different chromium runs. Libraries were pooled based on their molar concentrations and sequenced on an Illumina NovaSeq 6000 system using an S1 flow cell and v1 300 cycle Reagent kit with 28 cycles for read 1, 90 cycles for read 2, 10 cycles for index1 (i7) and 10 cycles for index2 (i5).
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
nuclear RNA
|
| Data processing |
Library strategy: snRNA-seq
Cellranger (4.0.0) mkfastq was used to perform sample demultiplexing and generate fastq files
Cellranger (4.0.0) count was used to generate the single nucleus gen-UMI counting. Reads were mapped to mm10 (GENCODE vM23/Ensembl 98, from 10x).
The resulting data matrix was analyzed uisng Seurat (v3). Doublets were removed from each GEM reaction using DoubletFinder. In addition to doublets, we removed nuclei with more than 1% expression of mitochondrial RNAs. Expression was log scale normalized and the top 200- features were used for PCA and downstream clustering and UMAP. Harmony was used to combine the separate repliates.
Clusters were called using FindClusters (Seruat) and were annotated manually using key lineage markers.
For visualization of expression on UMAP projections, RNA expression values were imputed using Markov Affinity-based Graph Imputation of cells (MAGIC).
Genome_build: mm10
Supplementary_files_format_and_content: hdf5 files contain gene expression counts on a per nucleus basis for each sample; values represent filtered droplets (i.e. empty droplets removed) derived from cellranger
Supplementary_files_format_and_content: barcode.tsv.gz files contains list of barcodes for each sample; for import into Seurat
Supplementary_files_format_and_content: features.tsv.gz files contains list of features (genes) for each sample; for import into Seurat
Supplementary_files_format_and_content: matrix.mtx.gz files contains values of each feature for each barcode for each sample; for import into Seurat
|
| |
|
| Submission date |
Jul 06, 2021 |
| Last update date |
Sep 14, 2021 |
| Contact name |
Fiorella Carla Grandi |
| Organization name |
Gladstone
|
| Street address |
1650 Owens Street
|
| City |
San Francisco |
| ZIP/Postal code |
94305 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE179541 |
Complement factor C1q mediates sleep spindle disruption and epileptic spikes after mild brain injury |
|
| Relations |
| BioSample |
SAMN20080808 |
| SRA |
SRX11361091 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5420222_tbi2.matrix.mtx.gz |
40.9 Mb |
(ftp)(http) |
MTX |
| GSM5420222_tbi2_barcodes.tsv.gz |
23.8 Kb |
(ftp)(http) |
TSV |
| GSM5420222_tbi2_features.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSM5420222_tbi2_filtered_feature_bc_matrix.h5 |
15.8 Mb |
(ftp)(http) |
H5 |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
|
|
|
|
 |
