|
| Status |
Public on Oct 22, 2020 |
| Title |
DIPG4_DMSO_3 |
| Sample type |
SRA |
| |
|
| Source name |
diffuse intrinsic pontine glioma
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: SU-DIPG04
passage: 10-15
agent: DMSO
|
| Treatment protocol |
The small-molecule inhibitor used in this study is PTC-028 (orb181369, Biorbyt). This drug was dissolved in DMSO to make stock concentrations to use in in vitro experiments.Stable knockdown of genes was carried out using shRNAs targeting different genes. Briefly, the cells (~100,000/well) were seeded onto a 6-well plate and the cells were transduced with the lentiviral shRNA using polybrene (polybrene final concentration of 8 ng/ml). Transduced cells were selected using puromycin (2µg/ml) and the cells were routinely maintained in the cell culture medium containing 1µg/ml of puromycin. ShRNAs targeting BMI1 was obtained from transOMIC technologies and from the Functional Genomic Core Facility at the University of Colorado, Denver.
|
| Growth protocol |
SU-DIPG04 cells were cultured in tumor stem media (TSM) consisting of all the following components from Invitrogen: Neurobasal-A medium (1:1), D-MEM/F-12 (1:1), HEPES Buffer Solution (10 mM), MEM Sodium Pyruvate Solution (1 mM), MEM Non-Essential Amino Acids (0.1 mM), GlutaMAX-I Supplement (1:100), Antibiotic-Antimycotic (1X), B-27 Supplement Minus Vitamin A (1X), except all the growth factors, human-basic FGF (20 ng/mL), human-EGF (20 ng/mL), human PDGF-AA (10 ng/mL), human PDGF-BB (10 ng/mL) from Shenandoah Biotech and the heparin (2 ug/mL) from StemCell Technologies.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA from cell lines were extracted with a RNeasy Plus RNA kit (Qiagen) and the purity of the RNA isolated was examined using an Agilent 2100 bioanalyzer.
mRNA libraries were made from the isolated pure RNA and RNA sequencing was performed using Nova-Seq.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
DIPG4_PTC028_raw_counts.txt
|
| Data processing |
The reads were trimmed and clipped for quality control in Trimmomatic (v0.36).
Read quality was checked for each sample using FastQC v0.10.1.
High quality reads were then imported into STAR (v2.4.0.1) for alignment into BAM files.
The function summarizeOverlaps from the GenomicAlignments package (v1.16.0) was used to count the reads from BAM files.
DESeq (v1.32.0) was then used to get differential expression genes on the raw counts.
Alignment was done using the transcriptome hg38 (from UCSC)
Genome_build: hg38
Supplementary_files_format_and_content: txt file includes raw read counts.
|
| |
|
| Submission date |
Jul 07, 2020 |
| Last update date |
Oct 22, 2020 |
| Contact name |
Etienne Danis |
| E-mail(s) |
etienne.danis@cuanschutz.edu
|
| Organization name |
University of Colorado, Denver - Anschutz Medical Campus
|
| Street address |
12800 E. 19th Ave
|
| City |
Aurora |
| State/province |
CO |
| ZIP/Postal code |
80045 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE140768 |
Senescence induced by BMI1 inhibition is a therapeutic vulnerability in H3K27M-mutant DIPG |
|
| Relations |
| BioSample |
SAMN15462426 |
| SRA |
SRX8677617 |
