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GEO help: Mouse over screen elements for information. |
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Status |
Public on Aug 21, 2023 |
Title |
Skin, excised, cultured, expanded, for Rep3 |
Sample type |
SRA |
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Source name |
Skin
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Organism |
Homo sapiens |
Characteristics |
tissue: Skin cell type: fibroblast genotype: WT treatment 1: hTRET-immorttalized treatment 2: untreated time: NA replicate: rep 3
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Treatment protocol |
For myo-conversion, immortalized fibroblasts were transduced with a Tet-on inducible lentiviral vector encoding mouse Myod with a multiplicity of injection of 10 in presence of 4 μg/ml of polybrene (Sigma-Aldrich). Cells were cultured in DMEM, 4.5 g/l glucose, 10% fetal bovine serum and 0.1% Penicil-lin-Streptomycin. Cells were then plated on dishes coated with 1% Matrigel (Sig-ma-Aldrich) at high density and cultured in DMEM, 10 µg/ml insulin and 4 µg/ml doxycycline (Sigma-Aldrich). After 5 days, myo-converted fibroblasts (McF) were harvest-ed for follow-up assays. For myogenic differentiation, cells were plated on dishes coated with 1% Matrigel (Sigma-Aldrich) at high density in differentiating medium (DMEM, 10 µg/ml insulin, 50 µg/ml Penicillin-Streptomycin). After 5 days, myotubes (MT) were harvested for follow-up assays.
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Growth protocol |
Immortalized fibroblasts were cultured in DMEM, 4.5 g/l glucose, 10% fetal bovine serum and 0.1% Penicillin-Streptomycin. Immortalized myoblasts were cultured in DMEM (Invitrogen) supplemented with 16% M199 (Invitrogen), 4.5 g/l glucose, 20% fetal bovine serum, 0.1% Penicillin-Streptomycin, 5 µg/ml insulin (Sigma-Aldrich), 25 µg/ml fetuin (Life Technologies), 5 ng/ml human Epidermal Growth Factor (Life Technologies), 0.5 ng/ml basic fibroblast growth factor (Life Technologies) and 0.2 µg/ml dexamethasone (Sigma-Aldrich).
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was extracted with TRIzol (Thermo Fisher Scientific), eluted in 20 µl RNase-free water and concentration estimated by A260. RNA integrity number was assessed with an Agilent 2100 Bioanalyzer using the Agilent RNA 6000 Nano kit Libraries were prepared using the KAPA mRNA HyperPrep kit (Roche)
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NovaSeq 6000 |
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Data processing |
RNA-seq reads were filtered to remove low-quality reads using fastp (v 0.23.2). Expression of transcripts was quantified using Salmon package36 (v1.7.0) and Ensembl GRCh38.p13 (release 108). Transcript abundance values were imported into R and summarized with tximport (v1.28.0 ) .DESeq2 (v1.36.0) as implemented in SARTools was then used to normalize raw counts and apply exploratory analysis.
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Submission date |
Jun 29, 2023 |
Last update date |
Aug 21, 2023 |
Contact name |
Philippe Collas |
Organization name |
University of Oslo
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Department |
Institute of Basic Medical Sciences
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Street address |
PO Box 1112 Blindern
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City |
Oslo |
ZIP/Postal code |
0317 |
Country |
Norway |
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Platform ID |
GPL24676 |
Series (2) |
GSE236118 |
A robust and practical myogenic cell system to explore cellular and genomic features of muscle differentiation [RNA-Seq] |
GSE236120 |
A robust and practical myogenic cell system to explore cellular and genomic features of muscle differentiation |
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Relations |
BioSample |
SAMN36071141 |
SRA |
SRX20824264 |
Supplementary file |
Size |
Download |
File type/resource |
GSM7518476_CTL_NI_3_S3_count.txt.gz |
223.0 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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