|
| Status |
Public on Jun 18, 2022 |
| Title |
MCF7-Y537S mutant cells + OTX015 [Y537S_OTX_1] |
| Sample type |
SRA |
| |
|
| Source name |
Breast
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Breast
cell line: MCF7
cell type: Epithelial cells
genotype: Y537S ESR1 mutation
treatment and_time: OTX015, 24h
|
| Treatment protocol |
Hormone depletion was carried over 3 days by washing the cells with phenol red-free IMEM without serum (5 washings daily, 1 hour apart). At completion of washings each day, cells were re-cultured in phenol red-free IMEM supplemented with 5% charcoal-dextran-treated calf serum. On day 4, the cells were trypsinized and 1 x 106 cells were seeded on 10-cm culture dish. After overnight culture, the cells were treated with DMSO, 10 pM β-estradiol, or 1μM OTX015 for 24 hours.
|
| Growth protocol |
MCF7 or MCF7-Y637S/D538G mutant cells were cultured in phenol red-free medium, IMEM supplemented with 5% charcoal-dextran-treated calf serum and 2% penicillin-streptomycin in hormone-depleted conditions in humidified atmosphere with 5% CO2 at 37C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The cells were then harvested and total RNA was extracted using RNeasy Mini Kit (QIAGEN) according to the manufacturer’s instructions. RNA quantity and purity was measured by a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific), and integrity was analyzed by an Agilent Bioanalyzer 2100 (Agilent Technologies).
RNA-seq libraries were prepared using Illumina TruSeq Standard mRNA Sample Preparation Kit (Illumina) according to the manufacturer’s protocol. Libraries were validated on an Agilent Bioanalyzer 2100. RNA-seq libraries were sequenced on a SE75 (single end 75 base pair) NextSeq500 flow cell.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Reads with phred quality scores less than 20 and less than 35 bp after trimming were removed from further analysis using trimgalore (v0.4.1). Quality-filtered reads were then aligned to the mouse reference genome GRCh38 (hg38) using the HISAT (v 2.0.1) aligner(41) using default settings and marked duplicates using Sambamba (v0.6.6)(42). Aligned reads were quantified using ‘featurecount’ (v1.4.6)(43) per gene ID against GENCODE v25.
Assembly: Quality-filtered reads were then aligned to the mouse reference genome GRCh38 (hg38) using the HISAT (v 2.0.1) aligner(41) using default settings and marked duplicates using Sambamba (v0.6.6)(42). Aligned reads were quantified using ‘featurecount’ (v1.4.6)(43) per gene ID against GENCODE v25.
Supplementary files format and content: Aligned reads were quantified using ‘featurecount’ (v1.4.6)(43) per gene ID against GENCODE v25.
|
| |
|
| Submission date |
Jun 15, 2022 |
| Last update date |
Jun 18, 2022 |
| Contact name |
SM N Udden |
| E-mail(s) |
[email protected]
|
| Organization name |
UT Southwestern Medical Center
|
| Department |
Radiation Oncology
|
| Street address |
2201 Inwood Road
|
| City |
Dallas |
| State/province |
TX |
| ZIP/Postal code |
75390-9187 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE206185 |
Targeting ESR1 mutation-induced transcriptional addiction with BET inhibition |
|
| Relations |
| BioSample |
SAMN29094304 |
| SRA |
SRX15712485 |
