2025-08-31
ENA first public
ERC000011
ENA-CHECKLIST
Erasmus MC
INSDC center name
public
INSDC status
ERS22559895
SRA accession
E-MTAB-14678:Library_5
Submitter Id
ArrayExpress
broker name
ESC derived cell line
cell_line
pool
cmo id
not collected
collection date
house mouse
common name
Protocols: Day 7 differentiated mouse embryonic stem cells are collected as embryonic bodies, washed with PBS, dissociated with TrypLE Express Enzyme for 5 minutes, and Trypsin-EDTA for 3 minutes at 37C in intermittent pipetting. After dissociation, samples are inactivated with PBS+10%FCs, go through a filter (40 um), and spin down 1000rpm for 5 minutes. The samples were washed three times with PBS+10%FCS. Than, 10X Genomics 3' CellPlex Kit Set A (CMO sample multiplexing) protocol followed and continued with  Chromium Next GEM Single Cell 3ʹReagent Kits v3.1. The harvested embryonic bodies were washed three times with PBS, and prewarmed TrypLE was added in a 50 ml falcon tube and placed in a 37°C water bath while shaking with intermittent pipetting for disassociation.  Four minutes later, 3 ml Trypsin/EDTA was added, and the dissociation process continued for at least 7 minutes while visually checking the dissociation of the EB. The disassociation buffer was inactivated with PBS+10%FCS, strained with a 40 µM cell strainer to create a single-cell suspension, washed three times, and stained for cell surface marker staining or prepared for scRNA-seq.   mESCs-Medium: DMEM HyClone™ (cytiva SH30081), 15% - Fetal Bovine Serum (FBS) (Capricorn Scientific FBS-12A, CP18-2152), 1X - MEM Non-Essential Amino Acids Solution (100X) (Gibco 11140-035), 2mM - GlutaMAX™ Supplement (Gibco 35050061), 10mM - HyClone™ HEPES Buffer (cytiva SH30237.01), 1X - Penicillin-Streptomycin (100X) (Sigma-Aldrich P0781), 0.1mM - 2-Mercaptoethanol (50 mM) (Gibco 31350010), 1000U.ml -  ESGRO® Recombinant Mouse LIF Protein 10^7 units/ml (Sigma-Aldrich ESG1107).    Differentiation Medium:  IMDM, GlutaMAX™ Supplement (Gibco 31980048), 15% - FBS (Capricorn Scientific FBS-12A, CP18-2152), 1X - Penicillin-Streptomycin (100X) (Sigma-Aldrich P0781), 0.1 mM - 2-Mercaptoethanol (50 mM) (Gibco 31350010), 50 µg/ml - L-Ascorbic acid (Sigma-Aldrich A5960-25G), 150 μg/mL -Transferrin (Roche 10652202001)  Chromium Next GEM Single Cell 3' Reagent Kits v3.1 protocol has been followed. Chromium Next GEM Single Cell 3' Reagent Kits v3.1 protocol has been followed with dual indexing library construction.
description
embryo stage
developmental stage
not collected
geographic location (country and/or sea)
multiplexed pool 1
individual
not applicable
isolate
Mus musculus
organism
Mus musculus
organism
embryoid body
organism part
Mus musculus
scientific_name
XY
sex
129X1/SvJ
strain
Library_5
title