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Status |
Public on May 09, 2020 |
Title |
Control-2 |
Sample type |
RNA |
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Source name |
hiPSC-derived DMD patient myoblasts
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Organism |
Homo sapiens |
Characteristics |
cell type: hiPSC-derived DMD patient myoblasts treatment: DMSO treated for 24 hours
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Treatment protocol |
Triplicate samples were used in microarray analysis. D2 hiPSC-derived myoblasts were treated with ginsenoside (5μM), fenofibrate (8μM) and DMSO for 24h in the differentiation medium.
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Growth protocol |
The DMD(D2325) hiPSC-derived myoblasts which carries mutation c.457 C->T were differentiated using the CHIR-DAPT protocol . Briefly, hiPSCs were plated as single cells on Geltrex (Gibco) treated dishes, at a density of 1.5 x 105 cells per well of a 24-well plate, in the presence of MEF-conditioned N2 media containing 10 ng/ml of FGF-2 (PeproTech) and 10 μM of Y-27632 (Cayman). The cells were induced to differentiate into myoblasts by adding CHIR99021 (3μM) in N2 medium for 4 days and by DAPT (10 μM) for the following 8 days. Afterwards cells continue to differentiate and mature in N2 medium for the next 13 days. Myoblasts were collected by FACS with the selection marker NCAM+/HNK1-. The NCAM+/HNK1- myoblasts were maintained in a humidified incubator containing 5% CO2 at 37°C, and grown in N2 media supplemented with 10% fetal bovine serum (FBS). To induce myotube formation, expanded NCAM+/HNK1- myoblasts were plated to confluence, and switched to N2 media without serum.
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Extracted molecule |
total RNA |
Extraction protocol |
RNA was extracted from Trizol-suspended cells with the QIAGEN RNeasy Mini Kit according to manufacturer's protocol.
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Label |
biotin
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Label protocol |
RNA was labeled with the Affymetrix GeneChip WT PLUS Reagent Kit using the manufacturer's protocol.
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Hybridization protocol |
17 hours at 45° C with rotation (60rpm) as described by Affymetrix in their GeneChip Expression Analysis Technical Manual.
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Scan protocol |
Using Affymetrix’ GeneChip Scanner 3000 7G and default parameters described by the manufacturer in their GeneChip Expression Analysis Technical Manual.
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Description |
D2325 hiPSC-derived myoblasts treated DMSO(control)-rep2
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Data processing |
Affymetrix CEL files were extracted with the Partek GS 6.6 platform and their data RMA and quantile-normalized to create log2 transcript signal values, which were used in subsequent ANOVA analysis.
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Submission date |
Oct 09, 2018 |
Last update date |
May 11, 2020 |
Contact name |
Gabsang Lee |
E-mail(s) |
glee48@jhmi.edu
|
Phone |
443-287-8631
|
Organization name |
The Johns Hopkins University
|
Department |
School of Medicine
|
Lab |
Institute for Cell Engineering
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Street address |
733 North Broadway
|
City |
Baltimore |
State/province |
MD |
ZIP/Postal code |
21205 |
Country |
USA |
|
|
Platform ID |
GPL23159 |
Series (1) |
GSE121023 |
Gene expression data from DMD patient hiPSC-derived myoblasts treated with fenofibrate, ginsenoside Rd or DMSO |
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