|
| Status |
Public on Mar 07, 2019 |
| Title |
MDA231-DTX-EV |
| Sample type |
SRA |
| |
|
| Source name |
Extracellular vesicles
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MDA-MB-231
treatment: docetaxel (DTX)
sample type: Extracellular vesicles
molecule subtype: miRNA
|
| Treatment protocol |
Cells were treated with docetaxel (DTX; 4 nM), doxorubicin (DOXO; 125 nM), or PBS for 48 h in growth medium containing EV-depleted FBS.
|
| Growth protocol |
MDA-MB-231 cells were grown in DMEM plus 10% fetal bovine serum (FBS). To collect extracellular vesicles (EV), conditioned medium was first prepared by incubating cells grown at sub-confluence in growth medium containing EV-depleted FBS (prepared by overnight ultracentrifugation of medium-diluted sera at 100,000 ×g at 4 °C) for 48 h, and pre-cleared by centrifugation at 500 ×g for 15 min and then at 10,000 ×g for 20 min. EVs were pelleted by ultracentrifugation at 110,000 ×g for 70 min, and washed in PBS under the same ultracentrifugation conditions.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
TRIZOL LS reagent (Thermo Fisher Scientific) was used to extract total RNA from EVs following the manufacturer’s protocol. RNA pellet was dissolved in RNase-free water, and subjected to library construction and deep sequencing.
Each sample was independently subjected to library preparation and deep sequencing. All small RNAs of 15–52 nts were selected and sequenced using the Illumina system, following the manufacturer’s protocol (Illumina Inc., San Diego, CA). Library preparation, as well as cluster generation and deep sequencing, was performed according to the 5' ligation-dependent (5′ monophosphate-dependent) manufacturer’s protocol (Digital Gene Expression for small RNA; Illumina). Small RNAs were size-selected between 17 and 52 nt according to the single-stranded DNA marker in the small RNA sequencing kit (Illumina). The library was quantified using picoGreen and qPCR. Sequencing was performed on a Genome Analyzer IIx (Illumina), and image processing and base calling were conducted using Illumina’s pipeline.
|
| |
|
| Library strategy |
miRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
11317-MDA231-EV-Docetaxel_TCATTC_L999
|
| Data processing |
Sequenced reads were adapter trimmed and mapped onto human genome version hg19 using Bowtie v1 software and the expression level of mature miRNAs in the miRBase human miRNA database v15 was summarized.
Normalization and identification of differentially expressed miRNAs between groups were carried out using Bioconductor package “edgeR”.
Genome_build: hg19
Supplementary_files_format_and_content: miRNA expression counts in text format
|
| |
|
| Submission date |
Feb 11, 2019 |
| Last update date |
Mar 07, 2019 |
| Contact name |
Xiwei Wu |
| E-mail(s) |
xwu@coh.org
|
| Organization name |
City of Hope National Medical Center
|
| Department |
Computational and Quantitative Medicine
|
| Street address |
1500 E. Duarte Rd.
|
| City |
Duarte |
| State/province |
CA |
| ZIP/Postal code |
91010 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE126419 |
Small RNA sequencing of extracellular vesicles from MDA-MB-231 cells treated with PBS, docetaxel (DTX), or doxorubicin (DOXO) |
|
| Relations |
| BioSample |
SAMN10914552 |
| SRA |
SRX5361504 |
