|
| Status |
Public on May 21, 2015 |
| Title |
IL2 knockout GM-DCs, replicate 2 |
| Sample type |
RNA |
| |
|
| Source name |
GM-DC derived from bone marrow and selected using CD11c microbeads
|
| Organism |
Mus musculus |
| Characteristics |
strain: B6.129P2-Il2tm1Hor/J
cell type: GM-CSF myeloid dendritic cells
genotype: IL2 knockout
|
| Treatment protocol |
Wt and IL-2-/- GM-DCs were treated or not with curdlan for 6h and processed for microarray
|
| Growth protocol |
Bone marrow was flushed with IMDM medium with GM-CSF (200ng/ml), 1% Penicillin-Streptomycin, 10% Serum, 1% L-Glutamine and 0.1% β-Mercaptoethanol and cells were cultured for 7d The suspension cells were harvested on d7 and purified by positive selection using CD11c microbeads (Miltenyi Biotec) according to the manufacturer’s protocol. GM-DC (CD11c+, MHCII aI-A/I-E +) purity determined by FACS was routinely >95%.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted following the double extraction protocol using Trizol and Qiagen RNeasy Micro Extraction Kit. Total RNA integrity was assessed by Agilent Bioanalyzer.
|
| Label |
biotin
|
| Label protocol |
Single-stranded DNA were prepared according to the Ambion WT Expression Kit from 100ng of total RNA. 5.5ug of ssDNA were fragmented and labeled using the Affymetrix GeneChip WT Terminal Labeling Kit.
|
| |
|
| Hybridization protocol |
Fragmented samples were hybridized according to Affymetrix guidelines for 17hr in 45°C on GeneChip Mouse Gene 1.0 ST Array; The arrays were then washed and stained using the standard fluidics protocol (FS450_0007) for gene arrays on the GeneChip Fluidics Station 450.
|
| Scan protocol |
The mouse gene chips were scanned using GeneChip Scanner 3000. The images were analyzed using Expression Console and standard array quality controls analyses were carried out according to the instructions provided by Affymetrix.
|
| Data processing |
CEL files were processed in Bioconductor using the xps package. Expression data was summarized at the transcript level and normalized using RMA. Log2 transformation was applied to the expression data.
|
| |
|
| Submission date |
May 30, 2014 |
| Last update date |
May 22, 2015 |
| Contact name |
Bernett Lee |
| E-mail(s) |
bernett_lee@immunol.a-star.edu.sg
|
| Organization name |
Singapore Immunology Network
|
| Street address |
8A Biomedical Grove #03-06 Immunos
|
| City |
NA |
| ZIP/Postal code |
138648 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE58120 |
Dendritic cell-derived IL-2 promotes apoptosis of terminally mature cells via a novel autocrine signaling pathway |
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