|
| Status |
Public on Jun 15, 2021 |
| Title |
19-1245_714R1: iEF + EF (714) Replicate 1 |
| Sample type |
SRA |
| |
|
| Source name |
A673 cells
|
| Organism |
Homo sapiens |
| Characteristics |
disease state of cells: Ewing sarcoma
cell line: A673
|
| Treatment protocol |
Endogenous EWS/FLI protein was knocked down in A673 cells using an shRNA targeting the 3'-UTR of the native transcript (iEF); iLuc (a shRNA targeting the firefly luciferase gene) was used as a control for knockdown. cDNA constructs were expressed: empty vector control (Empty Vector), EWS/FLI (EF), or two mutant EWS/FLI constructs (EF DBD and EF DBD+). Antiobiotic selection was used to select for cells expressing these constructs and cells were collected after 8 days. Whole cell total RNA was extracted using a Qiagen RNeasy Extraction kit and genomic DNA was removed uisng Qiagen gDNA Eliminator columns.
|
| Growth protocol |
Ewing sarcoma cells (A673) were seeded at 50-70% confluency and left to adhere overnight.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy kit (Qiagen) and gDNA Eliminator columns (Qiagen) were used to remove residual genomic DNA. 3 ug of total RNA was used for library prep.
TruSeq Stranded mRNA Kit
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
RNA-sequencing was performed on A673 cells with endogenous EWS/FLI knocked-down and EWS/FLI cDNA (EF(714)) was expressed.
|
| Data processing |
FASTQC and Multiqc were used for raw sequence quality control.
Trim_galore was used to trim low-quality and overrepresented sequences.
Two-pass STAR appraoch (STAR version 2.5.2b) was used to align sequences to hg19 human reference genome.
RSeQC was used to evaluate post-mapping sequence quality.
DESeq2 was used to identify different analysis of genes.
Genome_build: hg19
Supplementary_files_format_and_content: fastq files include raw reads of each samples; bigwig files are aligned, trimmed, normalized reads combined for each samples from separate replicates.
|
| |
|
| Submission date |
Nov 05, 2020 |
| Last update date |
Sep 30, 2021 |
| Contact name |
Megann A. Boone |
| E-mail(s) |
[email protected]
|
| Organization name |
Nationwide Children's Hospital
|
| Department |
Center for Childhood Cancer
|
| Lab |
Dr. Stephen Lessnick
|
| Street address |
700 Children's Drive, WA5011
|
| City |
Columbus |
| State/province |
OH |
| ZIP/Postal code |
43205 |
| Country |
USA |
| |
|
| Platform ID |
GPL20301 |
| Series (2) |
| GSE160897 |
The FLI portion of EWS/FLI contributes a transcriptional regulatory function that is distinct from its DNA-binding function in Ewing sarcoma [RNA-seq] |
| GSE160898 |
The FLI portion of EWS/FLI contributes a transcriptional regulatory function that is distinct from its DNA-binding function in Ewing sarcoma |
|
| Relations |
| Reanalyzed by |
GSE185130 |
| BioSample |
SAMN16675304 |
| SRA |
SRX9443560 |
