|
| Status |
Public on May 02, 2012 |
| Title |
Non-targeting shRNA rep 1 RNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
Non-targeting shRNA RNA-seq
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HEK293T
cell type: human embryonic kidney cells
|
| Treatment protocol |
HEK293T cells were transfected with AFF2, AFF3, or AFF4 SMARTpool siRNA duplex (Dharmacon) using Lipofectamine (Invitrogen). AFF2 shRNA (V2THS_113929), AFF3 shRNA (V2THS_133851), and AFF4 shRNA (V2THS_197522) and constructs were purchased from Open Biosystems. Lentiviral particle preparation and infection were performed as previously described (Lin et al. 2010). Total RNA was extracted with the RNeasy kit from Qiagen and residual DNA was digested with RNase-free DNase I (Sigma) before re-purification by the RNeasy kit.
|
| Growth protocol |
HEK293T cells were cultured in DMEM supplemented with 10% FBS. Cells were maintained at 37 °C under 5% CO2.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was prepared from the shRNA stable cell lines, and biological duplicate experiments were performed. Polyadenylated RNA was purified from total RNA and fragmented. Double-stranded cDNA from the RNA fragments were ligated to adapters and subjected to deep sequencing (Illumina GA IIx).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Description |
NT_shRNA_1
|
| Data processing |
NT-1.bam; genome build: hg19
RNA-seq analysis was done using TopHat v1.3.2 (Trapnell et al. 2009) and Bowtie v0.12.7. Human transcript annotations were from RefSeq (downloaded from UCSC on Jan 21, 2011). Differentially expressed genes after RNAi were called with a FDRadjusted p-value < 0.05 and MA plots were determined by Cufflinks v1.1.0 (Trapnell et al. 2010). Paired-end read fragment insert lengths were estimated at a mean of 205 with a standard deviation of 145. Four non-targeting RNAi samples were used as the control samples. Each RNAi sample was done with two replicates.
|
| |
|
| Submission date |
Dec 02, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Alexander (Garrett) Garruss |
| E-mail(s) |
asg@stowers.org
|
| Organization name |
Stowers Institute for Medical Research
|
| Lab |
Shilatifard
|
| Street address |
1000 East 50th Street
|
| City |
Kansas CIty |
| State/province |
Missouri |
| ZIP/Postal code |
64110 |
| Country |
USA |
| |
|
| Platform ID |
GPL10999 |
| Series (1) |
| GSE34097 |
The SEC family of RNA Polymerase II elongation factors: gene target specificity and transcriptional output |
|
| Relations |
| SRA |
SRX110220 |
| BioSample |
SAMN00761663 |
