|
| Status |
Public on Aug 18, 2020 |
| Title |
Human_436_20B_UNC_d4_3 |
| Sample type |
SRA |
| |
|
| Source name |
MDA-MB-436
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: TNBC
paclitaxel sensitivity: parental taxol-sensitive treated with paclitaxel 72h
treatment: UNC1999 3uM, 96h
replicate: GD_RNA_70 and 56
|
| Treatment protocol |
Taxol-resistant cells were grown in presence of 20nM of Paclitaxel at all time. Parental cells (MDA-MB-436 and Hs 578T) were grown in DMEM with 1:1000 DMSO at all time.
|
| Growth protocol |
MDA-MB-436 and HS 578T cells were grown in DMSO supplemented with 10%FBS and Penicilin/Streptomycin (100 U/mL Penicilium and 100 μg/mL. Streptomycin). Taxane-resistant cells were generated upon long-term exposure to increasing concentrations of paclitaxel at a starting concentration 0.05 nM with increments (dose and time adjusted to the growth and survival of adapting cells), until the cytotoxic concentration of 10 to 20 nM was reached with the resistant cells growing at a similar rate than the parental cells (>6 months).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the qiaquick purification kit
RNA samples were quantified by qubit (Life Technologies) and quality by Agilent Bioananlyzer. Two hundred nanograms Total RNA from 42 samples were library prepared using TruSeq Stranded Total RNA kit (Illumina). RNA samples were ribosomal RNA depleted using Ribo-zero Gold rRNA beads, following purification the RNA was fragmented. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using RNase H and DNA Polymerase I. A single “A” based were added and adapter ligated followed by purification and enrichment with PCR to create cDNA libraries.
Final cDNA libraries were size validated using Agilent Bioanalyzer and concentration validated by qPCR (Kapa Biosystems/Roche). All libraries were normalized to 10nM and pooled together, denatured with 0.2N NaOH and diluted to a final concentration of 1.4pM. 1.3ml of 1.4pM pooled libraries were loaded onto an Illumina NextSeq cartridge for cluster generation and sequencing on an Illumina Nextseq500 instrument (Illumina) using Paired-end 75bp protocol to achieve ~ 40 million reads per sample
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Reads were aligned to hg19 using tophat (2.0.8) with default parameters
Transcripts were merged using cufflinks (2.1.1) and differential expression computed for all relevant contrasts with default parameters
Genome_build: hg19
Supplementary_files_format_and_content: mean FPKM value for all identified transcripts
|
| |
|
| Submission date |
Mar 15, 2018 |
| Last update date |
Aug 18, 2020 |
| Contact name |
Alexander James Murison |
| E-mail(s) |
[email protected]
|
| Organization name |
University Health Network
|
| Lab |
Dick Lab
|
| Street address |
101 College Street, TMDT room 11-706
|
| City |
Toronto |
| State/province |
Ontario |
| ZIP/Postal code |
M5G 1L7 |
| Country |
Canada |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
| GSE111920 |
Metabolic adaptations underlie epigenetic vulnerabilities in chemoresistant breast cancer. [RNA-Seq1] |
| GSE113687 |
Metabolic adaptations underlie epigenetic vulnerabilities in chemoresistant breast cancer |
|
| Relations |
| BioSample |
SAMN08720346 |
| SRA |
SRX3800611 |
