|
| Status |
Public on Jul 16, 2019 |
| Title |
shYAP1 E2 GROseq |
| Sample type |
SRA |
| |
|
| Source name |
MCF7
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF7
cell type: Breast cancer cell line
shRNA: shYAP1
treatment: 100 nM E2, 1hr
|
| Treatment protocol |
For knockdown of TEAD4 and YAP1, MCF7 cells were infected with shRNA lentiviruses and selected by puromycin (1 ug/ml) for 3 days to establish TEAD4 or YAP knockdown MCF7 stable cell lines. Before experiment, the MCF7 cells were hormone-stripped in phenol red-free DMEM medium plus 5% charcoal-treated FBS for 3 days, followed by treatment of either 100 nM 17β-estradiol (E2) or ethanol as vehicle control for 1 hour.
|
| Growth protocol |
MCF7 obtained from ATCC were cultured in DMEM media supplemented with 10% FBS in a 5% CO2 humidified incubator at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
MCF7 cells were subjected to nuclear run-on for 5 minutes at 30°C with BrU labeling. The run-on RNAs were pulled down by BrU beads and subjected to library preparation for deep sequencing.
We followed a previous published protocol (Ingolia et al., 2009 Science; Wang et al., 2011 Nature). Briefly, the run-on RNA were first added with a polyA tracts by RNA polyA polymerase. This polyA tail enables the run-on RNA to be reverse transcribed into single strand cDNA by Reverse Transcriptase. The cDNA was circularized by CircLigase (Epicentre) and re-linearized by APE I, subsequently subjected to PCR amplification and deep sequencing.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 3000 |
| |
|
| Description |
shYAP1_E2_GROseq
GRO-seq, shYAP1, 1hr treatment with 100nM E2.
Nascent RNA
|
| Data processing |
Library strategy: GRO-seq
Basecalls performed using Illumina CASAVA software.
GRO-seq reads were aligned to human genome (hg19) using bowtie with “--best --strata –m 1 –v 2” parameters. Duplicated reads were eliminated for subsequent analysis.
To balance the clonal amplification bias and total useful reads, only at most three reads was allowed for each unique genomic position.
edgeR 3.16.5 was used to call the differential eRNAs.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: bigwig
|
| |
|
| Submission date |
Jan 24, 2019 |
| Last update date |
Jul 16, 2019 |
| Contact name |
Zhijie Jason Liu |
| E-mail(s) |
liuz7@uthscsa.edu
|
| Organization name |
Universality of Texas Health Science Center at San Antonio
|
| Department |
Department of Molecular Medicine
|
| Lab |
Liu lab
|
| Street address |
7703 Floyd Curl Drive
|
| City |
San Antonio |
| State/province |
TEXAS |
| ZIP/Postal code |
78229 |
| Country |
USA |
| |
|
| Platform ID |
GPL21290 |
| Series (2) |
| GSE125607 |
A non-canonical role of YAP/TEAD is required for activation of estrogen-regulated enhancers in breast cancer [GRO-seq] |
| GSE125609 |
A non-canonical role of YAP/TEAD is required for activation of estrogen-regulated enhancers in breast cancer |
|
| Relations |
| BioSample |
SAMN10814312 |
| SRA |
SRX5287879 |
