|
| Status |
Public on Mar 19, 2018 |
| Title |
mRNA_siC_rep1 |
| Sample type |
SRA |
| |
|
| Source name |
ovarian cancer derived cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: ES-2
cell type: ovarian cancer derived cell line
sirna: Control siRNA
|
| Treatment protocol |
Parental ES-2 cells were seeded for RNA extraction.
|
| Growth protocol |
ES-2 cells were cultured in DMEM supplemented with 10% FBS. Cells were grown at 37°C in 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction was performed using TRIZOL according to manufacturer's protocol.
500 ng of total RNA was depleted of ribosomal RNA using the Ribo-Zero rRNA removal kit (Epicentre) according to the instructions of the manufacturer. Depleted RNA was then fragmented by addition of 5 fragmentation buffer (200 mM Tris acetate, pH 8.2, 500 mM potassium acetate and 150 mM magnesium acetate) and heating at 94 °C for 3 min in a thermocycler followed by ethanol precipitation with ammonium acetate and GlycoBlue (Life Technologies) as carrier. Fragmented RNA was then reversed transcribed using random hexamer and Superscript III (Life Technologies). The second strand was synthesized using the TargetAmp kit (Epicentre) according to the instructions of the manufacturer. The final steps of library preparation, e.g. blunt end repair, adapter ligation, adapter fill-in and amplification were done according to Meyer and Kircher (2010, PMID: 20516186). The barcoded libraries were purified and quantified using the Library Quantification Kit - Illumina/Universal (KAPA Biosystems) according to the instructions of the manufacturer. A pool of up to 10 libraries was used for cluster generation at a concentration of 10nM using an Illumina cBot. Sequencing of 2x101 bp was performed with an IlluminaHighScan-SQ sequencer at the sequencing core facility of the IZKF Leipzig (Faculty of Medicine, University Leipzig) using version 3 chemistry and flowcell according to the instructions of the manufacturer.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiScanSQ |
| |
|
| Description |
NB_C1
|
| Data processing |
Sequenced reads were trimmed for adaptor sequences and low quality sequence using Cutadapt (v 1.6) with parameters -q 10 -O 7 -m 20
Trimmed reads were mapped against the human genome (hg19 UCSC) using TopHat2 (v 2.0.13) with parameters -p 6 -g 20 --b2-N 1
Mapped reads were summarized using featureCounts (v 1.4.6) with parameters -p -M -t exon -g gene_id and Ensembl gene annotations (GRCh 37.75)
Differential gene expression was assessed using R/edgeR (v 3.12) using TMM normalization
Genome_build: hg19
Supplementary_files_format_and_content: Comma-separated text file including FPM, log2 fold change and false discovery rate values for each sample
|
| |
|
| Submission date |
Jan 24, 2018 |
| Last update date |
Mar 19, 2018 |
| Contact name |
Markus Glaß |
| E-mail(s) |
[email protected]
|
| Organization name |
Martin Luther University Halle-Wittenberg
|
| Department |
Institute of Molecular Medicine
|
| Lab |
Huettelmaier Lab
|
| Street address |
Kurt-Mothes-Str. 3a
|
| City |
Halle |
| ZIP/Postal code |
06120 |
| Country |
Germany |
| |
|
| Platform ID |
GPL15456 |
| Series (2) |
| GSE109604 |
IGF2BP1 enhances an aggressive tumor cell phenotype by impairing miRNA-directed downregulation of oncogenic factors [mRNA] |
| GSE109605 |
IGF2BP1 enhances an aggressive tumor cell phenotype by impairing miRNA-directed downregulation of oncogenic factors |
|
| Relations |
| BioSample |
SAMN08391411 |
| SRA |
SRX3597961 |
