|
| Status |
Public on Jan 01, 2020 |
| Title |
MIGA2-GFP_Day6_2 |
| Sample type |
SRA |
| |
|
| Source name |
3T3-L1
|
| Organism |
Mus musculus |
| Characteristics |
cell type: 3T3-L1
differentiation stage: 6 days after differentiation
genotype/variation: overexpressed MIGA2-GFP
|
| Treatment protocol |
To start differentiation the cells were cultured until they reached complete confluence, and were then further incubated for another 48 h. After 48 h at 100 % confluence the medium was exchanged with differentiation medium, (DMEM with 10% FBS, 1 µM Dexamethasone, 0.5 mM IBMX and 1 µg/mL Insulin). The time point when differentiation medium was added is defined as the start of differentiation in our time course experiments (Day 0). After two days the differentiation medium was replaced by DMEM/ 10 % FBS containing insulin (1 µg/mL). At day 4 after start of differentiation the medium was exchanged with DMEM with 10 % FBS. At day 6 after start of differentiation the cells were harvested.
|
| Growth protocol |
3T3-L1 pre-adipocytes were purchased from American Type Culture Collection (ATCC). 3T3-L1 cells were cultured at 37°C in DMEM (Thermo-Fisher) containing 10 % calf serum (CS) (Sigma-Aldrich) in 5% CO2. The cells were cultured in 10 cm tissue culture dishes (TPP) and the medium was exchanged at least every 2 days. The pre-adipocytes never reached densities above 70% confluence unless they were subjected to differentiation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The RNA of cell extracts of an adipocyte time course experiment was purified using RNeasy Mini Kit (Quiagen).
TruSeq RNA stranded protocol
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
bcl2Fastq v2.19 software used for demultiplexing + basecalling.
reads trimmed with trimmomatic, options: ILLUMINACLIP:adapters.fa:1:20:7:5:true AVGQUAL:10 MINLEN:20
reads mapped with STAR v2.5.4b, options: --sjdbOverhang 150 --twopassMode None --runThreadN 8 --outFilterType BySJout --outFilterMatchNmin 30 --outFilterMismatchNmax 10 --outFilterMismatchNoverLmax 0.05 --outMultimapperOrder Random --alignSJDBoverhangMin 1 --alignSJoverhangMin 8 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outFilterMultimapNmax 50 --chimSegmentMin 15 --chimJunctionOverhangMin 15 --chimScoreMin 15 --chimScoreSeparation 10 --outSAMstrandField intronMotif --outSAMattributes All --outStd BAM_Unsorted --outSAMtype BAM Unsorted
count: FeatureCounts (Bioconductor), options: allowMultiOverlap = true; countPrimaryAlignmentsOnly= true; minFeatureOverlap=10; minMapQuality= 10 ; keepMultiHits=true
differential expression analysis: EdgeR, options: normMethod = TMM; backgroundExpression =10; testMethod = glm; deTest =QL
Genome_build: Mus_musculus, Ensembl GRCm38.p5, Release_91-2018-02-26
Supplementary_files_format_and_content: tab-delimited, raw counts
|
| |
|
| Submission date |
Aug 10, 2019 |
| Last update date |
Jan 02, 2020 |
| Contact name |
giancarlo russo |
| E-mail(s) |
giancarlo.russo@fgcz.ethz.ch
|
| Organization name |
ETH/University of Zurich
|
| Department |
Functional Genomics Center Zurich
|
| Street address |
winterthurerstrasse 190
|
| City |
Zurich |
| State/province |
Schweiz |
| ZIP/Postal code |
8057 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE135683 |
Transcriptome analysis of adipocyte differentiation (3T3-L1 WT and MIGA2 perturbed/overexpressed) |
|
| Relations |
| BioSample |
SAMN12548830 |
| SRA |
SRX6693273 |
