Mus musculus
Protocols: Primary MEFs were isolated from E13.5 animals and cultured in standard medium containing Dulbecco's modified Eagle medium (DMEM) supplemented with 10% Fetal Bovine Serum (FBS), 50μg/ml streptomycin, 50 U/ml penicillin (Sigma) and 2mM L glutamine (Gibco). Cells were rinsed with PBS, exposed to tRA (10 μM) (Sigma-Aldrich) and cultured at 37oC for 16h. Total RNA was isolated from cells using a Total RNA isolation kit (Qiagen) as described by the manufacturer. Libraries were prepared using the Illumina® TruSeq® mRNA stranded sample preparation kit. Library preparation started with 1ug total RNA. After poly-A selection (using poly-T oligo-attached magnetic beads), mRNA was purified and fragmented using divalent cations under elevated temperature. The RNA fragments underwent reverse transcription using random primers. This is followed by second strand cDNA synthesis with DNA Polymerase I and RNase H. After end repair and A-tailing, indexing adapters were ligated. The products were then purified and amplified (14 PCR cycles) to create the final cDNA libraries. After library validation and quantification (Agilent 2100 Bioanalyzer), equimolar amounts of library were pooled. The pool was quantified by using the Peqlab KAPA Library Quantification Kit and the Applied Biosystems 7900HT Sequence Detection System.
ENA-FIRST-PUBLIC
2020-03-25T04:04:01Z
ENA-LAST-UPDATE
2019-07-19T15:29:42Z
External Id
SAMEA5800012
INSDC center name
IMBB-FORTH
INSDC first public
2020-03-25T04:04:01Z
INSDC last update
2019-07-19T15:29:42Z
INSDC status
public
Submitter Id
E-MTAB-8156:WT Rep2
broker name
ArrayExpress
cell line
mouse embryonic fibroblast cell
cell type
fibroblast
common name
house mouse
developmental stage
E13.5
genotype
wild type genotype
organism part
embryo
sample name
E-MTAB-8156:WT Rep2
scientific_name
Mus musculus
strain
C57BL/6