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| Status |
Public on Aug 14, 2019 |
| Title |
O1_TA_Hi_14 [scRNA-seq] |
| Sample type |
SRA |
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| Source name |
Tibialis anterior muscles
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| Organism |
Mus musculus |
| Characteristics |
strain: MGI: 5308730
cell type: Muscle stem cells
age: 24 months
mouse id: O1
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| Treatment protocol |
Muscles were chopped and put into a 15 ml Falcon tube containing 10 ml of DMEM, 0.08% collagenase D (Sigma, 11 088 882 001), 0.1% trypsin (ThermoFisher, 15090), 10 µg/ml DNaseI (Sigma, 11284932) at 37°C under gentle agitation for 25 min. The digestion was repeated 3 times until complete digestion of the muscle. The supernatants were filtered through a 70-µm cell strainer (Miltenyi, 130-098-462). Cells were spun for 15 min at 515g at 4°C and the pellets were resuspended in 1 ml freezing medium (10% DMSO (Sigma, D2438) in foetal calf serum (FCS, Invitrogen)) for long term storage in liquid nitrogen. Before FACS isolation, samples were thawed in 50 ml of cold DMEM, spun for 15 min at 515g at 4°C. Pellets were resuspended in 300 µl of DMEM 2% FCS 1 µg/mL propidium iodide (Calbiochem, 537060) and filtered through a 40-µm cell strainer (BD Falcon, 352235). Viable cells were isolated based on size, granulosity and GFP expression level (top 10% nGFPHi cells) using a MoFlo Astrios cell sorter (Beckmann Coulter). Single cells were collected in 2.5 µL cold RLT Plus buffer (Qiagen, 1053393) containing 1U/µL RNase inhibitor (Ambion, AM2694) in 96 well-plates (LoBind Eppendorf, 0030129504), flash-frozen on dry ice and stored at -80°C.
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| Extracted molecule |
total RNA |
| Extraction protocol |
We prepared scM&T-seq libraries by isolating mRNA on magnetic beads and separating from the single-cell lysate prior to reverse transcription and amplification using Smartseq2 but with 25 PCR cycles.
scM&T-seq (C. Angermueller et al., 2016)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
mRNA
processed data file: Table_raw_counts_RNA_QSC.txt
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| Data processing |
Libraries were sequenced on the Illumina HiSeq platform using the default RTA analysis software.
Genome_build: GRCm38
Supplementary_files_format_and_content: Table_raw_counts_RNA_QSC.txt file contains the raw counts per gene quantified over the exons using the RNA-seq quantitation pipeline in Seqmonk software (www.bioinformatics.babraham.ac.uk/projects/seqmonk/) in the format: <gene name> <chromosome> <start> <counts cell1> <counts cell 2> .... < counts celln>
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| Submission date |
Oct 16, 2018 |
| Last update date |
Aug 14, 2019 |
| Contact name |
Felix Krueger |
| E-mail(s) |
fkrueger@altoslabs.com
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| Organization name |
Altos Labs
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| Department |
Bioinformatics
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| Street address |
Granta Park
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| City |
Cambridge |
| ZIP/Postal code |
CB21 6GP |
| Country |
United Kingdom |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE121364 |
Ageing affects DNA methylation and transcriptional cell-to-cell variability in muscle stem cells [scRNA-seq] |
| GSE121437 |
Ageing affects DNA methylation and transcriptional cell-to-cell variability in muscle stem cells |
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| Relations |
| BioSample |
SAMN10249427 |
| SRA |
SRX4895199 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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