|
| Status |
Public on May 31, 2023 |
| Title |
JB_67 [RNA-seq] |
| Sample type |
SRA |
| |
|
| Source name |
primary neuroblastoma tumor
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: primary neuroblastoma tumor
tumor stage: 4S
mycn status: not amplified
|
| Extracted molecule |
total RNA |
| Extraction protocol |
500 ng of total RNA were depleted of ribosomal RNA using the RiboMinus kit (Life Technologies) 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 reverse 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 2x100 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 |
Neuroblastoma_tumor_totalRNA_TMM_FPKM.csv
DE_MNA_nMNA_protein_coding.csv
DE_17_unbalanced_protein_coding.csv
|
| Data processing |
Sequenced strand unspecific paired-end reads were trimmed for adaptor sequences and low quality sequence using Cutadapt (v 1.18) with parameters -q 20 -O 7 -m 20 –trim-n
Trimmed reads were mapped against the human genome (hg38 UCSC) using HiSat2 (v 2.1.0) with parameters -q --dta -k 5
Secondary alignments were filtered out using samtools (v 1.10).
Mapped reads were summarized using featureCounts (v 2.0.0) with parameterst -p -s 0 -T 6 -M -t exon -g gene_id and Ensembl gene annotations (GRCh 38.96)
Differential gene expression was assessed using R/edgeR (v 4.0.3) using TMM normalization.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: Comma-separated text files including FPKM, log2 fold change and false discovery rate values for each sample.
|
| |
|
| Submission date |
Aug 06, 2021 |
| Last update date |
May 31, 2023 |
| Contact name |
Danny Misiak |
| E-mail(s) |
danny.misiak@medizin.uni-halle.de
|
| Phone |
+49345573962
|
| Organization name |
Martin Luther University Halle-Wittenberg
|
| Department |
Molecular Cell Biology
|
| Lab |
Hüttelmaier Lab
|
| Street address |
Kurt-Mothes-Str. 3a
|
| City |
Halle (Saale) |
| ZIP/Postal code |
06120 |
| Country |
Germany |
| |
|
| Platform ID |
GPL15456 |
| Series (2) |
| GSE181559 |
IGF2BP1 induces neuroblastoma via a druggable feedforward loop with MYCN promoting 17q oncogene expression [human RNA-seq] |
| GSE181582 |
IGF2BP1 induces neuroblastoma via a druggable feedforward loop with MYCN promoting 17q oncogene expression |
|
| Relations |
| BioSample |
SAMN20601033 |
| SRA |
SRX11661483 |
