|
| Status |
Public on Nov 04, 2015 |
| Title |
U2AF2 RIP U2AF1 siRNA 3 [exon-level] |
| Sample type |
RNA |
| |
|
| Source name |
U2AF2 RIP from U2AF1 siRNA treatment of activated CD4 T cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Primary CD4 T cells
pretreated with: U2AF1 siRNA for 24hrs
activated with: anti-CD3/CD26 activator beads for 24 h
passages: 7-8
rip antibody: U2AF2 antibody
rip antibody vendor: Sigma
rip antibody cat. #: U4758
rip antibody lot #: 034K4841
molecule subtype: U2AF2 RIP RNA
|
| Treatment protocol |
Primary CD4 T cells cultured were treated with siRNA 24 hours prior to treatment with anti-CD3/CD26 activator beads (Thermo) for an additional 24 hours.
|
| Growth protocol |
Primary CD4 T cells cultured for two weeks in RPMI supplemented with 10% FBS, 30U/ml IL2, 100U/ml penicillin and 100mg/ml streptomycin at 37C and 5% CO2
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cell pellets were resuspended in TRIZOL reagent (Thermo) and the RNA was extracted according to manufacturer's protocol. RNA resuspended in nanopure water was purified over an Rneasy column (QIAGEN) and the polyA RNA was selected using oligo dT beads.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cDNA was prepared using the Ambion WT Expression Kit (Ambion) according to the manufacturer's instructions. cDNA was framented and labeled with the GeneChip WT Terminal Labeling Kit (Affymetrix).
|
| |
|
| Hybridization protocol |
Standard Affymetrix 49 Format Hybridization with 15mg of fragmented cRNA
|
| Scan protocol |
Affymetrix EukGE-WS2v4 on GCS 3000 7G
|
| Description |
SAMPLE 6
|
| Data processing |
Expression intenisties at the probe set (exon) and transcript (gene) level were made with Expression Console (Affymetrix).
Gene level expression intensities were used as input into the R Bioconductor package limma for identification of differentially expressed genes (U2AF2 RIP Ctrl siRNA vs. U2AF2 RIP Specific siRNA)
Exon level expression intensitied were used as input into Alt-Analyze (Emig, et al) to identify differentially spliced exons (U2AFA2 RIP Ctrl siRNA vs. U2AF2 RIP Specific siRNA)
Please note that the 'U2AF2RIP_HTA2_all_siRNA_100414.xlsx' file includes multiple worksheets containing: 1) Differentially expressed genes in ctrl siRNA vs. U2AF1 siRNA; 2)Differntially spliced genes in ctrl siRNA vs. U2AF1 siRNA; 3)Differentially spliced exons in ctrl siRNA vs. U2AF1 siRNA; 4)Differentially expressed genes in ctrl siRNA vs. SYNCRIP siRNA; 5)Differntially spliced genes in ctrl siRNA vs. SYNCRIP siRNA; 6)Differentially spliced exons in ctrl siRNA vs. SYNCRIP siRNA
probe group file: HuEx-1_0-st-v2.r2.pgf
meta-probeset file: HuEx-1_0-st-v2.r2.dt1.hg18.full.mps
|
| |
|
| Submission date |
Nov 03, 2014 |
| Last update date |
Nov 04, 2015 |
| Contact name |
Thomas C Whisenant |
| E-mail(s) |
[email protected]
|
| Organization name |
The Scripps Research Institute
|
| Department |
Molecular and Experimental Medicine
|
| Lab |
Salomon
|
| Street address |
10550 North Torrey Pines Rd
|
| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92037 |
| Country |
USA |
| |
|
| Platform ID |
GPL17585 |
| Series (2) |
| GSE62922 |
The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells [RIP-array] |
| GSE62923 |
The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells |
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