|
Status |
Public on Mar 07, 2016 |
Title |
SRSF4_iCLIP_mm9_Rep3 |
Sample type |
SRA |
|
|
Source name |
P19
|
Organism |
Mus musculus |
Characteristics |
cell line: P19 embryonal carcinoma cell line fusion protein expression: SRSF4-GFP antibody: anti-EGFP (D. Drechsel, MPI-CBG) barcode sequence: GGTC
|
Treatment protocol |
Cells were washed with PBS and irradiated with 254 nm UV light to crosslink proteins to nucleic acids.
|
Growth protocol |
Cells were grown in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal calf serum and pennicilin/streptomycin antibiotics at 37˚C with 5% CO2.
|
Extracted molecule |
total RNA |
Extraction protocol |
Cells were lysed and treated with RNase I. GFP-tagged proteins were immunoprecipitated with Gamma binding beads coupled to goat anti-EGFP antibody (D. Drechsel, MPI-CBG). After stringent washing, RNA was dephosphorylated and ligated to a 3' linker. Protein-RNA complexes were resolved by denaturing gel electrophoresis and transfered to nitrocellulose membrane. A membrane region above the expected size of the protein was cut and RNA was released by Proteinase K treatment. RNA was reverse transcribed using reverse primer containing a barcode sequence. cDNA was separated into 3 size fractions by denaturing gel electrophoresis and circularized by ssDNA circligase. A specific oligonukelotide was annealed to create a restriction site for BamHI and cDNA was linearized by digest with BamHI. Linearized cDNA was PCR amplifiied with Solexa sequencing primers to create the final libraries. For more details see: König et al. (2010): iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959
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|
|
Library strategy |
RIP-Seq |
Library source |
transcriptomic |
Library selection |
other |
Instrument model |
Illumina HiSeq 2000 |
|
|
Description |
processed data file: mSRSF4_iCLIP_all_mm9_iCLIP_tags.bed processed data file: mSRSF4_iCLIP_all_mm9_iCLIP_tag_clusters.bed
|
Data processing |
Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. (2010): iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome_build: mm9 Supplementary_files_format_and_content: .bed files containing the genomic coordinates for significant CLIP tags for individual replicates or combined replicates (3-6 replicates). Supplementary_files_format_and_content: .bed files containing the genomic coordinates for significant CLIP tag clusters from combined replicates.
|
|
|
Submission date |
Jun 09, 2015 |
Last update date |
May 15, 2019 |
Contact name |
Michaela Müller-McNicoll |
E-mail(s) |
[email protected]
|
Phone |
+49 69 798 42079
|
Organization name |
Goethe University Frankfurt
|
Department |
Institute for Cell Biology and Neuroscience
|
Lab |
RNA Regulation Group
|
Street address |
Max-von-Laue-Str. 13
|
City |
Frankfurt am Main |
ZIP/Postal code |
60438 |
Country |
Germany |
|
|
Platform ID |
GPL13112 |
Series (2) |
GSE69689 |
Transcriptome-wide mapping of RNA:protein interactions of seven SR proteins and NXF1 in P19 cells by iCLIP |
GSE69734 |
SR protein family in P19 cells |
|
Relations |
BioSample |
SAMN03766052 |
SRA |
SRX1054350 |