|
| Status |
Public on Dec 15, 2011 |
| Title |
S469_KO45_2 |
| Sample type |
RNA |
| |
|
| Source name |
bone marrow derived macrophages
|
| Organism |
Mus musculus |
| Characteristics |
cell type: bone marrow cells
strain: C57Bl/6
genotype: TTP-/-
time after actd: 45 min
rep: 2
|
| Treatment protocol |
BMDMs from Wt and TTP-/- mice were treated with LPS for 3 h. Medium was then replaced by fresh medium containing actinomycin D and SB203580 for 0, 45 and 90 min before RNA preparation.
|
| Growth protocol |
Bone marrow cells from C57Bl/6 mice were differentiated to macrophages (BMDM) with M-CSF-containing medium
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with Trizol (Invitrogen Life Technologies, Carlsbad, CA) according to the manufacturer’s instruction. Concentration and integrity of total RNA were determined by spectrophotometry (Nanodrop) and Experion automated electrophoresis system (Biorad, Munich, Germany).
|
| Label |
biotin
|
| Label protocol |
RNA samples were processed and hybridised to Affymetrix Mouse Gene ST 1.0 GeneChips according to the manufacturer’s protocols. In brief, 200 ng total RNA and 100 ng nascent RNA were reverse transcribed introducing by random priming a T7-binding site into the cDNA that allows in vitro transcription. The resulting cRNA was subjected to a second round of random primed cDNA synthesis in the presence of dUTP, that allows fragmentation of the cDNA with uracil DNA glycosylase and apurinic/apyrimidinic endonuclease 1. Biotinylation of the fragmented cDNA was accomplished by incubation with Terminal Deoxynucleotidyltransferase (TdT).
|
| |
|
| Hybridization protocol |
5 µg of biotinylated DNA were hybridized to Mouse Gene ST 1.0 GeneChips overnight, followed by washing and staining procedures, and scanning, following Affymetrix protocols.
|
| Scan protocol |
Scanning following Affymetrix protocols
|
| Description |
Gene expression data from bone marrow derived macrophages
|
| Data processing |
For generation of probe set expression values, CEL files containing probe level data were normalized using RMA (Affymetrix Expression Console).
|
| |
|
| Submission date |
Apr 27, 2011 |
| Last update date |
Jun 03, 2013 |
| Contact name |
Roland Lang |
| E-mail(s) |
[email protected]
|
| Organization name |
University Hospital Erlangen
|
| Street address |
Wasserturmstr. 3-5
|
| City |
Erlangen |
| ZIP/Postal code |
91054 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE28880 |
TTP-dependent mRNA decay in LPS-stimulated macrophages |
|
