|
| Status |
Public on Jul 21, 2020 |
| Title |
HeLa_shINTS11_Proseq_dox_r2 |
| Sample type |
SRA |
| |
|
| Source name |
HeLa
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa
cell type: cervical adenocarcinoma
modification: Tet-pLKO_shINTS11 stable cells
|
| Biomaterial provider |
ATCC#CCL-2
|
| Growth protocol |
HeLa stable containing INTS11 shRNA cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS and puromycin (2 μg/ml). Knockdowns were induced by adding of 1 μg/ml oxycycline (Dox) into the culture medium daily for 3 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
PRO-seq experiments were performed as described previously (Mahat, D. B. et al. 2016). Nuclei were isolated by Dounce homogenizer with loose pestle. 1x107 nuclei were subjected to nuclear run-on (30oC, 3min) in the presence of 25 μM Biotin-11-ATP/GTP/CTP/UTP (PerkinElmer). Total RNA was extracted and hydrolyzed in 0.2 M NaOH (on ice, 10 min).
PRO-seq libraries used biotinylated nascent RNAs were purified by Dynabeads M-280 streptavidin (Invitrogen). Following adaptor ligation as described in previously (Mahat, D. B. et al. 2016), cDNA synthesis, and PCR amplification 140–350 bp libraries were size-selected by Pippin HT with 2% gel cassette 20B (Sage Science).
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
shINTS11
|
| Data processing |
Library strategy: PRO-seq
Proseq: FASTQ data were trimmed by Cutadapt 1.14 and Trimmomatic v0.32.
Proseq: Reads were aligned on hg19 or dm3 genome by bowtie 1.1.2 26. Strand-specific single nucleotide ends of aligned reads were generated by Bedtools v2.26 with genomecov as bedgraph format. Bedgraph data were normalized by the number of reads mapped to spike-in dm3 genome, and then converted to bigwig files.
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format: Proseq: bigwigs were with Bedtools v2.26 with genomecov.
|
| |
|
| Submission date |
Jan 23, 2019 |
| Last update date |
Jul 21, 2020 |
| Contact name |
Felipe Beckedorff |
| E-mail(s) |
beckedorff@gmail.com
|
| Organization name |
University of Miami
|
| Street address |
1501 NW 10th Avenue room 731.G , Miami, FL
|
| City |
Miami |
| State/province |
FL |
| ZIP/Postal code |
33136 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE125535 |
The Human Integrator Complex Facilitates Transcriptional Elongation by Endonucleolytic Cleavage of Nascent Transcripts |
|
| Relations |
| BioSample |
SAMN10794320 |
| SRA |
SRX5283847 |
