MeDIP-2
Mus musculus
Protocols: Cells were passaged every 2-3 days to maintain cultures between 10% and 90% confluency. For analysis, near-confluent cultures were released from culture vessels with trypsin, spun down, and snap-frozen in liquid nitrogen before further processing. E14tg2a mouse embryonic stem cells were grown in dishes pre-coated with 0.1 % gelatin (w/vol, Sigma) in GMEM supplemented with 10% fetal bovine serum, sodium pyruvate, 50 μM β-mercaptoethanol, glutamax, non-essential amino acids (all from Gibco/ThermoFisher) and 10 ng/ml murine leukemia inhibitory factor (LIF, Protein Expression Facility, MPI Dortmund). gDNA was isolated from E12tg2a mESCs using the Monarch Genomic DNA Purification Kit (NEB, T3010) according to the manufacturer's instructions. gDNA was sheared to an average fragment size of 200 bp using a Bioruptor Pico sonication device (Diagenode) For each immunoprecipitation, 1 µg of sheared gDNA was used as input. DIPs were performed according to the MagMeDIP-seq Package V2 protocol (Diagenode, C02010041), using either the provided anti-5mC antibody for MeDIP or 0.6 µg per IP of the mouse monoclonal anti-hmC antibody (Diagenode, C15200200) for hMeDIP. Additionally, a control IP with 0.6 µg of mouse IgG (Diagenode, C15400001) was included. Illumina libraries were prepared according to the Diagenode MeDIP-seq library preparation protocol. Final library concentrations were quantified using the Quantus Fluorometer (Promega) and fragment size distributions were assessed with a TapeStation (Agilent Technologies) using a D1000 ScreenTape. Libraries were sequenced on an Illumina NovaSeq X-25B instrument using paired-end 2x 150 bp reads with a target depth of 50 million reads per sample.
ENA first public
2026-04-14
INSDC center name
Max-Planck-Institute for Molecular Physiology
INSDC status
public
Submitter Id
E-MTAB-15961:MeDIP-2
broker name
ArrayExpress
cell line
ES-E14 cell
cell type
mouse embryonic stem cell
collection date
not collected
common name
house mouse
developmental stage
mouse embryo stage
genotype
wild type genotype
geographic location (country and/or sea)
not collected
isolate
not applicable
sample name
E-MTAB-15961:MeDIP-2
scientific_name
Mus musculus