25/EBMcells278
Mus musculus
Protocols: Cells were trypsinized with 0.5 % trypsin for 10 minutes at 37°C and collected by centrifugation at 300 g. Cell pellets were snap-frozen in liquid nitrogen and stored at -80°C until RNA extraction.  Postnatal day 1 mice were quickly decapitated and brains were extracted in ice-cold dissection medium (DMEM + 1 mM HEPES). Cortical hemispheres were dissected, followed by removal of cortical meninges and hippocampi using tweezers. Cortical tissues of 4 mice were pooled and minced into 2-3 mm sized pieces which were digested in 0.25 % trypsin for 20 min at 37°C. Trypsination was stopped by 5 washes with pre-warmed dissection medium, followed by tissue dissociation using sterile Pasteur pipettes of two consecutively decreasing tip diameters. After complete dissociation of the tissue, the cell suspension was diluted in 30 mL of DMEM supplemented with 10 % heat-inactivated fetal bovine serum (growth medium) and divided into two tissue culture grade T75 flasks pre-coated with 0.00075% w/v poly-D-lysine. Cells were allowed to reach confluency after about one week, and split at a 1:3 ratio into fresh, uncoated T75 flasks. Experiment was initated after passage 2. Cells were propagated for 2 passages in Dulbecco's modified Eagle's medium (DMEM) containing 10 % heat-inactivated fetal bovine serum at 37 °C and 5 % CO2.  100,000 mixed glial cells were plated on different growth surfaces in each well of a 6-well plate. PDL – rigid cell culture plastic surface coated with 0.00075 % w/v poly-D-lysine; PDL scratched – rigid cell culture plastic surface coated with 0.00075 % w/v poly-D-lysine, and the next morning, multiple scratches were applied to injure the confluent cell layer; Rigid OHA/GEL – rigid cell culture plastic surface pre-coated with 0.00075 % w/v poly-D-lysine, followed by coating with a mix of 1.25 % oxidized hyaluronic acid (OHA) and 2.5 % gelatin (GEL); 650 Pa OHA/GEL hydrogel – rigid cell culture plastic surface pre-coated with 0.00075 % w/v poly-D-lysine, followed by a mix of 5.0 % w/v oxidized hyaluronic acid and 2.5 % w/v gelatin and 0.75 % w/v microbial transglutaminase (mTG) for gelation by crosslinking; 250 Pa OHA/GEL hydrogel – rigid cell culture plastic surface pre-coated with 0.00075 % w/v poly-D-lysine, followed by a mix of 1.25 % w/v oxidized hyaluronic acid and 2.5 % w/v gelatin and 0.75 % w/v mTG for gelation by crosslinking. After plating, cells were allowed to attach overnight. On the next morning, medium was changed from DMEM + 10 % FBS (growth medium) to DMEM + 0.5 % FBS (assay medium). For the PDL scratched condition, the confluent cell layer was injured by multiple scratches applied with a 200 µL Eppendorf pipette plastic tip. Cells were allowed to grow in assay medium for 48 hours before cell harvest by trypsination. Total RNA from the cells was isolated from snap-frozen cell pellets using the RNeasy mini kit (Qiagen) with the application of an on-column DNase digestion step. Total RNA was eluted using 35 µL sterile, RNase-free water and the eluate was frozen until further processing for RNA sequencing. cDNA Libraries were constructed using the Illumina stranded mRNA kit.
ENA first public
2026-06-29
INSDC center name
FAU Erlangen Institute of Human Genetics, Universitätsklinikum Erlangen, FAU Erlangen-Nürnberg, Erlangen, Germany
INSDC status
public
Submitter Id
E-MTAB-17287:25/EBMcells278
age
1
broker name
ArrayExpress
collection date
not collected
common name
house mouse
developmental stage
Newborn (p1)
genotype
wild type genotype
geographic location (country and/or sea)
not collected
growth condition
650Pa_2d_diff
individual
pooled
isolate
not applicable
organism part
Neocortex (forebrain without hippocampus)
replicate
AstroM2
sample name
E-MTAB-17287:25/EBMcells278
scientific_name
Mus musculus
sex
Unknown
strain
C57BL/6J