|
| Status |
Public on Apr 25, 2019 |
| Title |
mESC - Rb TKO (D1.2) |
| Sample type |
SRA |
| |
|
| Source name |
mESC - Rb TKO
|
| Organism |
Mus musculus |
| Characteristics |
cell type: ESC
genotype: Rb TKO
|
| Growth protocol |
mESC lines were maintained feeder-free on 0.2% gelatin-coated (Sigma G9382) plates in Dulbecco’s modified Eagle’s medium (Gibco SH30243.01) containing 15% fetal bovine serum (Hyclone SH30071.03; VWR Seradigm 97068-085), MEM non-essential amino acids (Gibco 11140-050), penicillin-streptomycin-glutamine (Gibco 10378-016), and 0.1 mM beta-mercaptoethanol (Gibco 21985-023), and supplemented with LIF. Cells were passaged enzymatically using 0.05% Trypsin-EDTA (Gibco 15400-054). Single-cell suspensions of 1000 mESCs/mL were cultured in low density assays.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were grown at low density for four days. RNA extraction was performed using the RNeasy micro kit (Qiagen 74004).
Libraries were prepared using the TruSeq Stranded mRNA Library Preparation Kit (Illumina RS-122-2101).
Four wild-type and four E2F4KO mESC lines were sequenced on a HiSeq4000, generating on average 42 million paired-end (2x76) reads per sample. Two of the wild-type lines (J3.3 and J3.14) were expanded from clones that had retained E2F4 protein expression following transfection with a gRNA against E2F4, and the other two wild-type lines (C1 and C2) were obtained following transfection of an empty pX330 vector. Similarly, four TKO and four QKO lines were sequenced on a separate run of a HiSeq4000, also generating on average 45 million paired-end (2x76) reads per sample.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
D1.2
|
| Data processing |
Reads were mapped to mouse reference genome mm10 with STAR2.5.1b using default settings. Genes that have at least ten reads in two out of four samples of either the control group or the E2F4KO group were further used for analysis. Differentially expressed genes were obtained using DEseq2
Genome_build: mm10
Supplementary_files_format_and_content: File including counts and FPM, log2 Foldchanges and p-values obtained with DEseq2
|
| |
|
| Submission date |
Jan 25, 2018 |
| Last update date |
Apr 25, 2019 |
| Contact name |
Julia Arand |
| E-mail(s) |
juliaarand@googlemail.com
|
| Organization name |
Medical University of Vienna
|
| Street address |
Schwarzspanierstr. 16
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL21103 |
| Series (1) |
| GSE109684 |
E2F4 regulates transcriptional activation in mouse embryonic stem cells independently of the RB family |
|
| Relations |
| BioSample |
SAMN08397776 |
| SRA |
SRX3601333 |
