|
| Status |
Public on Jun 13, 2017 |
| Title |
shPML |
| Sample type |
SRA |
| |
|
| Source name |
U2OS Cell line
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: U2OS osteosarcoma cell line
genotype/variation: PML silencing
|
| Growth protocol |
Low passage U2OS osteosarcoma cells (source: ATCC, HTB-96) containing lentivirus-delivered shRNAs targeting luciferase (shControl, GTGCGTTGCTAGTACCAAC) or exon 4 of PML (shPML, AGATGCAGCTGTATCCAAG) were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) and 1µg/ml of puromycin at 37°C and 5% CO2. Cells were grown to 80% confluence in 10cm dishes prior to solubilization with Trizol.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with Trizol and RNA quality was assessed with the Agilent Bioanalyzer 2100.
PolyA-purified mRNA was subsequently used for cDNA-library construction with the Illumina TruSeq RNA Preparation Kit. Strand specificity was performed with 2'-deoxyuridine 5'-triphosphate during second-strand synthesis. All samples were indexed with Illumina adapters and each cDNA library was sequenced on an Illumina HiSeq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiScanSQ |
| |
|
| Description |
U2OS shPML
|
| Data processing |
Sequencing reads were mapped using TopHat (version 2.0.0) to the known transcriptome (-G option; Gencode version 7 annotations) for the human reference genome (hg19) using default parameters (Trapnell et al. 2009).
Cufflinks (version 2.0.2) was used to quantify gene and isoform expression for known transcripts (-G option; Gencode version 7 annotations) using the default parameters (Trapnell et al. 2010).
Only uniquely aligned reads were used for expression quantification. Pairwise differential gene expression analysis was performed using Cuffdiff (Trapnell et al 2012) with the default parameters.
Genome_build: hg19
Supplementary_files_format_and_content: Cufflinks output for gene and transcript FPKM
|
| |
|
| Submission date |
Jan 26, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Stephen B Montgomery |
| E-mail(s) |
smontgom@stanford.edu
|
| Organization name |
Stanford University
|
| Department |
Pathology
|
| Street address |
300 Pasteur Dr
|
| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305-5324 |
| Country |
USA |
| |
|
| Platform ID |
GPL15456 |
| Series (1) |
| GSE94126 |
Next Generation Sequencing of Gene expression changes in U2OS osteosarcoma cells with PML silencing |
|
| Relations |
| BioSample |
SAMN06275667 |
| SRA |
SRX2521842 |
