|
| Status |
Public on Aug 07, 2024 |
| Title |
DNA methylation array - Day 5 iPSC control Rep1_BS |
| Sample type |
genomic |
| |
|
| Source name |
Day 5 iPSC
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Day 5 iPSC
treatment: D5 control
|
| Treatment protocol |
Mouse recombinant IFNγ (10 ng/ml) (R&D Systems, 485-MI-100) was added to the reprogramming medium from day 0 to day 5.
|
| Growth protocol |
Reprogramming was induced in the presence or absence of IFNγ (from day 0 to 5). SSEA1+ day 5 iPSCs (4 replicates from different reprogramming rounds) and SSEA1+ X-GFP+ day 7 iPSCs (2 replicates from different reprogramming rounds) were sorted by FACS.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted and purified from reprogramming sorted cells using Qiagen DNeasy Kit according to standard instructions
|
| Label |
Cy5 and Cy3
|
| Label protocol |
Cy5 and Cy3
|
| |
|
| Hybridization protocol |
1µg of each sample was evenly splitted for the oxidation reaction and the mock-oxidation reaction where the oxidant solution was replaced by water following the TrueMethyl oxBS Module manufacturer’s instructions (NuGEN-Tecan, 0414). Both aliquots were then processed in parallel for all stages of the protocol. After the oxidation reaction where 5-hydroxymethylcytosine is oxidized to 5-formylcytosine (5fC) and 5-methylcytosine (5mC) stays unchanged, the bisulfite treatment converts 5fC and all non-methylated cytosines to uracil, while 5mC is not altered. For samples to be run on the Illumina Infinium® Mouse Methylation BeadChip Array (Illumina, 20041558), 7 μL of recovered TrueMethyl template were mixed with 1 μL of 0.4 N NaOH following manufacturer’s instructions. All subsequent steps were completed following the Infinium HD Assay Methylation protocol (https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/chemistry_documentation/infinium_assays/infinium_hd_methylation/infinium-hd-methylation-guide-15019519-01.pdf).
|
| Scan protocol |
Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
|
| Data processing |
IDAT files were preprocessed and analyzed using the minfi R package (v1.42.0). Probes were filtered for a detection p-value threshold of 0.01, and cross-reactive probes were removed. The remaining data was normalized using the preprocessQuantile method. Limma R package was used to do an experimental batch-correction.
Normalized beta values; normalization was done for array intensity using the preprocessQuantile method and then an experimental batch-correction was done using the Limma R package
|
| |
|
| Submission date |
Jun 30, 2023 |
| Last update date |
Aug 07, 2024 |
| Contact name |
Mercedes Barrero |
| Organization name |
Centre for Genomic Regulation
|
| Street address |
Dr. Aiguader, 88
|
| City |
Barcelona |
| ZIP/Postal code |
08003 |
| Country |
Spain |
| |
|
| Platform ID |
GPL31950 |
| Series (2) |
| GSE236246 |
DNA methylation arrays: The Interferon γ Pathway Enhances Pluripotency and X-Chromosome Reactivation in iPSC Reprogramming |
| GSE236247 |
The Interferon γ Pathway Enhances Pluripotency and X-Chromosome Reactivation in iPSC Reprogramming |
|
