SAMN36652777
External Id
University of Connecticut Health Center
INSDC center name
2023-08-31T00:51:36.243Z
INSDC first public
2023-08-31T00:51:36.243Z
INSDC last update
SRS18374272
INSDC secondary accession
live
INSDC status
Model organism or animal
NCBI submission model
Model.organism.animal.1.0
NCBI submission package
SRS18374272
SRA accession
missing
collection_date
DIV 7
dev_stage
Wild type
genotype
missing
geo_loc_name
mESC (E14TG2a) cells between passages number 5 – 15 were routinely maintained in mESC medium (DMEM supplemented with 1x Glutamax, FBS, β-mercaptoethanol, MEM non-essential aminoacids, sodium pyruvate, and LIF) on MEF feeder or on gelatin-coated tissue culture dishes. mESC media was replaced daily and split every other day. mESC differentiation is performed following this protocol: 3.5x106 cells were plated onto 90 mm bacteriological dishes in 15 mL NPC medium (DMEM with L-glutamine supplemented with 10% FBS, 1X non-essential amino acids, and 550 μM β-mercaptoethanol). Media change was performed on day 2 (equivalent of neurons DIV -8). On day 4, 6, and 8, media change was performed and 5 μM of retinoic acid was added. On day 10, NPC aggregates were collected and dissociated with 1 mL of TrypLE at 37°C for 5-7 min. To halt the reaction, 8 mL of Trypsin inhibitor was added. The NPC aggregates were gently dissociated by pipetting up and down and filtered through 40 μm cell strainer. Cell suspension was diluted in N2 media (Neurobasal supplemented with 1X N2 and 2 mM glutamine) at 3x105 cells/mL. 10 mL of cells were plated onto PDL coated 100 mm cell culture dishes. Complete media change was performed at 4h (day 10) and 24h (day 11) with N2 media. On day 12 and 14 media was replaced with B27 media (Neurobasal supplemented with 1X B27 and 2 mM glutamine). Cells were maintained until day 17 (DIV 7) and then collected for RNA extraction.
growth_protocol
Mus musculus
http://purl.obolibrary.org/obo/NCBITaxon_10090
organism
male
sex
E14TG2a
strain
embryonic stem cell derived neurons
tissue
Model organism or animal sample from Mus musculus
title