|
| Status |
Public on Oct 16, 2011 |
| Title |
Mock-transduced mouse CD8+ T cells, pooled sample#3 |
| Sample type |
RNA |
| |
|
| Source name |
CD8+ T cells
|
| Organism |
Mus musculus |
| Characteristics |
treatment: Mock-transduced
batch: Array batch 1
strain: C57BL/6
|
| Treatment protocol |
Twenty four hours after activation, cells were transduced with pMSGV1-ID32aThy1.1 (ID3-transduced group) or pMSGV1-Thy1.1 (mock-transduced group) retrovirus following standard protocol. On day 5, 3e5 cells were transferred into C57BL/6 mice in conjunction with 2E7 pfu rvvhgp100 virus. Five days later, Thy1.1+CD8+ cells were sorted out from CD8+ enriched splenocytes and analyzed using microarrays.
|
| Growth protocol |
On day zero, transgenic mouse Pmel-1 CD8+ cells expressing a TCR specific for gp100, were enriched with a MACS negative selection kit (Miltenyi Biotech). Cells were activated on plates coated with 2 g/ml anti-CD3 antibody and 1 g/ml of soluble anti-CD28 antibody in culture medium containing 10 ng/ml of IL-2 (Chiron Corporation).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNEasy Micro kit (Qiagen), following manufatcturer's instructions
|
| Label |
biotin
|
| Label protocol |
WT expression kit (Ambion), WT Terminal Labeling Kit (Affymetrix), according to the manufatcturer's instructions
|
| |
|
| Hybridization protocol |
Samples were hybridized to WT Human Gene 1.0 ST arrays using Affymetrix hybridization kit materials, according to the manufacturrer's instructions. Sample cocktails were heated at 99°C for 5 minutes, then 45°C for 5 minutes, centrifuged at max speed for 1 minute. Eighty ul of hyb solution was transferred to each array, and hybridized 17 hours at 45°C at 60rpm. Fluidics washing protocol used was FS450_0007.
|
| Scan protocol |
Affymetrix Gene ChIP Scanner 3000 7G
|
| Description |
Pmel-1 mouse CD8+ T cells transduced with pMSGV1-Thy1.1, pooled sample derived from four mice, sample #3
|
| Data processing |
Data from .CEL files were imported into Partek Genomincs Suite using the RMA method. For gene level analyses, probeset level data were merged using the median probeset expression level.
probe group file: MoGene-1_0-st-v1.r4.pgf
meta-probeset file: MoGene-1_0-st-v1.r4.mps
|
| |
|
| Submission date |
Aug 11, 2010 |
| Last update date |
Oct 16, 2011 |
| Contact name |
Zoltan Pos |
| E-mail(s) |
pos.zoltan@outlook.com.com
|
| Phone |
+(36)12102930-56435
|
| Organization name |
Semmelweis University
|
| Department |
Dept. of Genetics, Cell- and Immunobiology
|
| Street address |
4 Nagyvarad ter
|
| City |
Budapest |
| ZIP/Postal code |
H-1089 |
| Country |
Hungary |
| |
|
| Platform ID |
GPL10740 |
| Series (1) |
| GSE23568 |
Expression data analyzing ID3 (Inhibitor of DNA binding 3)-affected mouse T cells |
|
| Relations |
| Alternative to |
GSM578144 (gene-level analysis) |
