characteristics: ERC000011
characteristics: ENA-CHECKLIST
characteristics: 2022-12-01T00:19:38Z
characteristics: ENA-FIRST-PUBLIC
characteristics: 2022-12-01T00:19:38Z
characteristics: ENA-LAST-UPDATE
characteristics: SAMEA10379640
characteristics: External Id
characteristics: Max Planck Institute for Biology of Ageing
characteristics: INSDC center name
characteristics: 2022-12-01T00:19:38Z
characteristics: INSDC first public
characteristics: 2022-12-01T00:19:38Z
characteristics: INSDC last update
characteristics: public
characteristics: INSDC status
characteristics: ERS8033197
characteristics: SRA accession
characteristics: E-MTAB-11060:Young_set_batch2_1
characteristics: Submitter Id
characteristics: 3
characteristics: month
characteristics: age
characteristics: Batch2
characteristics: batch
characteristics: ArrayExpress
characteristics: broker name
characteristics: house mouse
characteristics: common name
characteristics: Protocols: Organoid cultures of different passages (4-13) were transferred into medium supplemented with 10% matrigel and maintained in a 37°C incubator, 5% CO2, 20% O2 shaking at 80-90 rpm. The organoids were passaged every 4 days with mechanical dissociation of the matrigel. For the differentiation assays, the medium was exchanged with differentiation medium (Broutier et al., 2016) (Advanced DMEM F12, 10mM Hepes, 1%GlutaMAX, 1:50 B27 supplement (without vitamin A), 1 mM N-acetylcysteine, 10 nM recombinant human [Leu15]-gastrin I, 50 ng/ml recombinant mouse EGF, 100 ng/ml recombinant human FGF10, 50 nM A83-01 and 10 μM DAPT) after 3 days of culture and was re-newed every 2 days. The 13th-15th day of differentiation, the medium was changed daily with the supplementation of 3uM of dexamethasone. Assessment of differentiation was performed on day 16th. Total RNA from young and old organoids at passages 10-13 was extracted in two batches (batch1: n=4 and batch2: n=3 per age) with the Zymoresearch Direct-zol RNA Miniprep (R2051) kit. 50ng RNA per sample were used for cDNA synthesis with Maxima H Minus reverse transcriptase (Thermo Fisher Scientific). During reverse transcription, unique barcodes including unique molecular identifiers (UMI) were attached to each sample. After cDNA synthesis, all samples were pooled and processed in one single tube. DNA was purified using AmpureXP beads (Beckman Coulter) and the eluted cDNA was subjected to Exonuclease I treatment (New England Biolabs). cDNA was PCR-amplified for 12 cycles and subsequently purified. After purification, cDNA was tagmented in 10 technical replicates of 1 ng cDNA each using the Nextera XT Kit (Illumina), according to the manufacturer's instructions. The final library was purified and concentration and size were validated by Qubit and High Sensitivity TapeStation D1000 analyses.
characteristics: description
characteristics: organoid culture
characteristics: growth condition
characteristics: Mus musculus
characteristics: organism
characteristics: Mus musculus
characteristics: organism
characteristics: liver
characteristics: organism part
characteristics: 1
characteristics: replicate
characteristics: Mus musculus
characteristics: scientific_name
characteristics: Young_set_batch2_1
characteristics: title