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| Status |
Public on Apr 10, 2018 |
| Title |
1772117101_B09 |
| Sample type |
SRA |
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| Source name |
spinal cord dorsal horn
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| Organism |
Mus musculus |
| Characteristics |
age: 21
Sex: M
total molecules: 23061
cluster name: Glut_Reln_Nmur2
celltype: Glut8
strain: C57bl6
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| Extracted molecule |
total RNA |
| Extraction protocol |
Spinal cords of 3-4 week mice were collected in oxygenated artificial cerebral spinal fluid (ACSF) on ice. Grey matter dorsal horn was isolated and transferred into a dish with 37°C digestion solution [300µl TrypLE™ Express (Life Technologies; cat#12605-010), 2 ml Papain solution (Worthington Biochemical; cat#LK003178; 25U/ml in ACSF), 100µl DNAse I (Worthington Biochemical; cat#LK003172; 1mM in ACSF) and 100µl ACSF]. The tissue was triturated using glass Pasteur pipettes with decreasing diameter until cells became visible at the bottom of the plastic dish. The solution was filtered using a 30µm cell strainer and diluted with 2.4ml ACSF and 0.6ml Neurobasal-A medium (Gibco; cat#10888) and centrifuged at 100g for 4min at 4°C. The supernatant was removed and the pellet resuspended in 0.5ml ASCF and 0.5ml Neurobasal-A medium, supplemented with L-Glutamine (Gibco; cat#25030-123), B27 (Gibco; cat#17504-044) and Penicillin/Streptomycin (Sigma 50x; cat#P4458). The cell suspension was transferred onto an Optiprep gradient [85µl of Optiprep Density Solution (Sigma; cat#D1556) mixed with 457.5µl ACSF and 457.5µl Neurobasal media including supplements], and centrifuged at 80g for 10min at 4°C. The supernatant was removed until only 100-200µl remained. 10µl of DNaseI was added to avoid cell clustering. The single cell density was evaluated and adjusted to 500 cells per µl using ACSF for dilution.
1000 cells/μl were resuspended in artificial cerebrospinal fluid and added to the Reverse transcription mix to aim for sampling of around 3500 cells. All downstream cDNA synthesis (13 PCR cycles), library preparation and sequencing were carried
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
molecule_counts.xlsx
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| Data processing |
Processing was performed with the 10X Chromium cellranger pipline version 1.1 using default settings
The Unique Molecular Identifier and cellular barcode corresponding to each read have been appended to the read id separated by an underscore.
Genome_build: UCSC mm10
Supplementary_files_format_and_content: Excel file of mRNA molecule counts per gene per cell
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| Submission date |
Sep 13, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Sten Linnarsson |
| Organization name |
Karolinska Institutet
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| Department |
Medical Biochemistry and Biophysics
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| Lab |
Molecular Neurobiology
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| Street address |
Scheeles väg 1
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| City |
Stockholm |
| ZIP/Postal code |
171 65 |
| Country |
Sweden |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE103840 |
Single-cell RNA sequencing of sensory neurons in the mouse dorsal horn |
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| Relations |
| BioSample |
SAMN07639479 |
| SRA |
SRX3183819 |
| Supplementary data files not provided |
SRA Run Selector |
| Processed data are available on Series record |
| Raw data are available in SRA |
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