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Sample GSM7518476 Query DataSets for GSM7518476
Status Public on Aug 21, 2023
Title Skin, excised, cultured, expanded, for Rep3
Sample type SRA
 
Source name Skin
Organism Homo sapiens
Characteristics tissue: Skin
cell type: fibroblast
genotype: WT
treatment 1: hTRET-immorttalized
treatment 2: untreated
time: NA
replicate: rep 3
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.
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).
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)
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
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.
 
Submission date Jun 29, 2023
Last update date Aug 21, 2023
Contact name Philippe Collas
Organization name University of Oslo
Department Institute of Basic Medical Sciences
Street address PO Box 1112 Blindern
City Oslo
ZIP/Postal code 0317
Country Norway
 
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
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 SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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