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| Status |
Public on Mar 01, 2023 |
| Title |
d0-si-cdkn-r3 |
| Sample type |
SRA |
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| Source name |
hiPSC-BAP 3D cells, siCDKN2A D0
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| Organism |
Homo sapiens |
| Characteristics |
cell line: hiPSC-BAP
cell type: Human induced pluripotent stem cells differentiated into brown adipocytes progenitors
genotype: CDKN2A knockdown
treatment: adipogenic differentiation
time: Day 0
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| Treatment protocol |
To generate spheroids, a suspension of hiPSC-BAPs at 0.5~1x 106 cells/ml was prepared in growth medium, and 1ml of the cell suspension was seeded in one well of 24 well Ultra-Low Attachment (ULA) plate (Corning 3473). The plate was maintained for 3 days to generate spheroids. Then, to differentiate spheroids, the growth medium was changed to differentiation medium composed of EBM-2 (Lonza) supplemented with 0.1% FCS, IBMX (0.5 mM), dexamethasone (0.25 μM), T3 (0.2 nM), insulin (170 nM), rosiglitazone (1 μM), SB431542 (5 μM), and EGM-2 cocktail (Lonza, CC-3121) including ascorbic acid, hydrocortisone and EGF. IBMX and dexamethasone were maintained only for the first 3 days of differentiation. SB431542 and EGF were removed after the first 9 days of differentiation. Spheroids were maintained in differentiation medium up to 20 to 30 days, with medium changed once a week.
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| Growth protocol |
hiPSC-BAPs, derived from the hiPSC line NOK6 as previously described (Mohsen-Kanson et al., 2014) were grown in standard tissue culture conditions at 37C with 5% CO2. The growth medium is DMEM low glucose supplemented with L-glutamine (2mM), penicillin–streptomycin 5,000 IU/ml–5,000 g/ml (Pen/Strep), 10% FBS and 2.5 ng/ml FGF2.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from hiPSC-BAP 3D at D0 and D10 of differentiation using TRIzolTM Reagent (Sigma).
The libraries were sequenced on the NextSeq system (Illumina), using a paired-end 2x75 bp protocol.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
the demultiplexing of sequence data was performed using bcl2fastq Conversion Software (Illumina; bcl2fastq v2.19.1). Trimming of adapter sequences was performed using cutadapt software (version 1.7.1). Reads quality control was assessed using FastQC (v0.11.5). Subsequently, sequence reads from FASTQ files were aligned to the human genome GRCh38, downloaded from Ensembl. Alignement was performed using STAR aligner (version 2.5.2b). Over 19 million of 75 base pairs PE-reads reads were generated per sample.The normalized counts of the different genes and isoforms was performed using RSEM (v1.2.31) using a GTF from Ensembl 89. Finally differential expression was performed using R version 3.6.3 and DESeq2 package v1.24.0.
Assembly: the human genome GRCh38
Supplementary files format and content: Excel file include Differential expression DE_D0_D10
Supplementary files format and content: Excel file include Differential expression DE_Ctrl_Cdkn_D0
Supplementary files format and content: Excel file include Differential expression DE_Ctrl_Cdkn_D10
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| Submission date |
Jan 19, 2023 |
| Last update date |
Mar 03, 2023 |
| Contact name |
souhila amanzougarene |
| Organization name |
CNRS
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| Lab |
EGID
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| Street address |
1 place de Verdun
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| City |
LILLE |
| ZIP/Postal code |
59000 |
| Country |
France |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE223241 |
Knocking down CDKN2A in 3D hiPSC-derived brown adipose progenitors potentiates differentiation, oxydative metabolism and browning process |
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| Relations |
| BioSample |
SAMN32793674 |
| SRA |
SRX19069646 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6943790_d0-si-cdkn-r3.genes.results.gz |
1.4 Mb |
(ftp)(http) |
RESULTS |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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