|
| Status |
Public on Sep 03, 2013 |
| Title |
PAR-CLIP1 |
| Sample type |
SRA |
| |
|
| Source name |
PAR-CLIP1
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: human embryonic kidney (HEK) 293 T-REx Flp-In cell lines
antibody: monoclonal anti-FLAG (Sigma, F1804)
rna subset: RBM10 bound RNA
|
| Treatment protocol |
siRNA transfections were performed in antibiotics free medium, i.e. DMEM with 10% final concentration FBS, using RNAiMAX (Invitrogen). RBM10 overexpression was induced with 10 ng/mL doxycycline for 16 hours
|
| Growth protocol |
The HEK293 T-REx Flp-In cells were cultured in a 37 °C incubator with 5 % CO2 and maintainence medium containing: High glucose DMEM (Gibco) supplemented with 10% final concentration FBS (heat inactivated, Gibco), 100 U/ml of penicillin and 100 μg/ml streptomycin (Gibco), 15 μg/ml blasticidin (InvivoGen) and 100 μg/ml zeocin (InvivoGen). The stable cell lines were maintained in selective medium containing: DMEM with 10% final concentration of FBS, 100 U/ml of penicillin and 100 μg/ml streptomycin, 100 μg/ml hygromycin and 15 μg/ml blasticidin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For PAR-CLIP, RBM10-RNA complexes were revovered from the gel followed by protein digestion and phenol choloroform extaction.
PAR-CLIP seqeuncing libraries were constructed using original small RNA library preparation protocols. Briefly, the 3' and 5' adaptors were ligated to RNA separately. The ligated product was reverse transcribed and amplified to generate the sequencing library.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Library strategy: PAR-CLIP
PAR-CLIP reads were aligned to the human genome allowing at most one mismatch, or indel of one nucleotide. Uniquely mapped reads were overlapped to define binding clusters. For each cluster, the rbm10 binding site was defined as the site with the highest number of T to C conversions.
processed data files format and content: The .bed files contain RBM10 binding sites. Scores given in column 5 are the numbers of reads per binding cluster.
Base calling was done on the Illumina HiSeq 2000 machine using RTA 1.12. We kept only reads passing the filtering.
Genome_build: hg19
|
| |
|
| Submission date |
Mar 08, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Andreas Gogol-Döring |
| E-mail(s) |
andreas.gogol-doering@mni.thm.de
|
| Organization name |
Technische Hochschule MIttelhessen
|
| Department |
MNI
|
| Street address |
Wiesenstraße 14
|
| City |
Gießen |
| ZIP/Postal code |
35390 |
| Country |
Germany |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE44976 |
Global analysis of alternative splicing regulated by RBM10 |
|
| Relations |
| SRA |
SRX248571 |
| BioSample |
SAMN01974961 |
