|
| Status |
Public on Jun 30, 2024 |
| Title |
RNA-seq_NB1-M16_SPH |
| Sample type |
SRA |
| |
|
| Source name |
Spheroids
|
| Organism |
Homo sapiens |
| Characteristics |
patient: NB1
pdxs: M16
cell type: Spheroids
|
| Growth protocol |
The tumor samples were subjected to mechanical and enzymatic dissociation using Tumor Dissociation Kit (Miltenyi Biotec). The dissociated tumor cells were suspended in DMEM/F12 medium supplemented with 1% penicillin-streptomycin, 2% B27 (Invitrogen), and 20ng/ml of human recombinant FGF and EGF (Peprotech). Adherent cells were cultured in DMEM (catalog no. Gibco) supplemented with 10% of FBS (PAN-Biotech) and 1% of penicillin-streptomycin (Gibco). Cells were maintained at 37°C in humidified, 5% CO2 chambers. Spheroid was grown in ultralow attachment plates (Corning).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with miRCURY kit (Exiqon) using Lysis buffer + B-mercaptoethanol. For tumor samples, dissociation was performed following gentleMACS Dissociator kit instructions for soft tumors (Miltenyi Biotech).
Minimum 100 ng used for preparing sequencing libraries using the TruSeq mRNA stranded kit (Illumina) and sequencing was performed on a Hi-Seq Illumina Genome Analyzer (100 bp single-read).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
RNA-seq reads were mapped to the hg38 genome (only chromosomes 1 to 22, X, Y and M) using hisat2 (version 2.0.5) and the Ensembl gene annotations Homo sapiens GRCh38.87.
Read density tracks were generated using bedtools genomecov (version 2.29.0 option -split) for each genomic position and read counts were normalized to one million reads in the library and visualized using the IGV browser.
Raw read counts and RPKM values were calculated over gene exons using htseq-count (version 0.9.1). Differentially expressed genes were identified using DESeq2 (version 1.26.0) using the following thresholds: adjusted p-value <= 0.001 and fold change >= 2.
Gene ontology annotations were used to calculate the enrichment of biological processes and associated hypergeometric p-values of genes in each class compared to all genes.
Assembly: hg38
Supplementary files format and content: bigWig
|
| |
|
| Submission date |
Jan 27, 2023 |
| Last update date |
Jun 30, 2024 |
| Contact name |
Anais Flore Bardet |
| Organization name |
IGBMC
|
| Street address |
1 rue Laurent Fries
|
| City |
Illkirch |
| ZIP/Postal code |
67404 |
| Country |
France |
| |
|
| Platform ID |
GPL16791 |
| Series (2) |
| GSE223900 |
Pre-clinical spheroid models identify BMX as a therapeutic target for metastatic MYCN non-amplified neuroblastoma (RNA-Seq) |
| GSE223902 |
Pre-clinical spheroid models identify BMX as a therapeutic target for metastatic MYCN non-amplified neuroblastoma |
|
