2025-08-01
ENA first public
ERC000011
ENA-CHECKLIST
University of Cambridge,  Cambridge Stem Cell Institute
INSDC center name
public
INSDC status
ERS23999599
SRA accession
E-MTAB-15041:30min_dTAG_D
Submitter Id
ArrayExpress
broker name
embryonic stem cell
cell type
not collected
collection date
house mouse
common name
Protocols: Cells were cultured on 10cm plates. Cells from all conditions were harvested at the same time using accutase and spun down for 3 min, 600 x g in a swinging bucket rotor at room temperature. Cells were washed in PBS and resuspended in ice-cold NE1 buffer (20mM HEPES-KOH pH7.9, 10mM KCl, 0.5mM spermidine, 0.1% Triton X-100, 5% Glycerol, protease inhibitors) with gentle vortexing. After 10 minutes incubation on ice, the pellet was collected by centrifugation for 4 minutes at 1300 x g at 4C in a swinging bucket rotor. Nuclei was washed with 1X PBS before being resuspended in wash buffer with 10% DMSO to a concentration of 1 million cells per mL. Nuclei was aliquoted in cryotubes and frozen overnight at -80C in Mr. Frosty containers. ES cells were cultured on 0.1% gelatin-coated plates in 2i/LIF conditions. For ADNP depletion, 0.5mM dTAG (1:1000 from DMSO stock) was resuspended in pre-warmed culture media and transferred to the cells. WT cells were not treated with dTAG. DNA was extracted using Zymo DNA Clean and Concentrator Kit-5 (Cat. No. D4014). Barcoding for library prep was performed by PCR amplification using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs, Cat. No. 10169146) and NEBNext® index primers for Illumina sequencing.
description
Chd4-mAID Adnp-FKBP
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Mus musculus
organism
Mus musculus
organism
2-B
replicate
Mus musculus
scientific_name
30min_dTAG_D
title