|
| Status |
Public on Sep 23, 2019 |
| Title |
B10 strain CD4 splenocyte at T0, biological rep1 |
| Sample type |
RNA |
| |
|
| Source name |
B10 strain_CD4 splenocyte at T0
|
| Organism |
Mus musculus |
| Characteristics |
strain: B10
genotype/variation: B10Br
age: 8 Weeks
gender: female
tissue: CD4+ splenocytes.
time point: basal state (T0)
|
| Treatment protocol |
These cells were divided into non-stimulated or phorbol 12-myristate 13-acetate (10 ng ml-1), Boeringer Mannheim) and ionomycin (1uM, Sigma-Aldrich, Missouri)--stimulated (PMA/I) groups and cultured at 37◦C and 5% CO2 for 4 hours.
|
| Growth protocol |
Splenocytes where extracted and sorted using AutoMACS (Miltenyi Biotech) CD4 Beads (LT34 Miltenyi Biotech) and maintained in MLC supplemented with 10% fetal calf serum (CSL, Parkville, Victoria, Australia) 4 mM L--glutamine, 1 mM sodium pyruvate, 10 mM HEPES, 50 uM beta-mercaptoethanol, penicillin, streptomycin and neomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed followed by clean ups with RNeasy Mini Kits (Qiagen) according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from total RNA. Labelling performed by The Ramaciotti Centre at The University of New South Wales (Sydney, Australia).
|
| |
|
| Hybridization protocol |
Standard Affymetrix protocols followed. Hybridisation performed by The Ramaciotti Centre at The University of New South Wales (Sydney, Australia).
|
| Scan protocol |
GeneChips were scanned using the Affymetrix GeneChip Scanner 3000 high resolution scanner. Scanning performed by The Ramaciotti Centre at The University of New South Wales (Sydney, Australia).
|
| Description |
HF_B1N
Gene expression data from B10 strain unstimulated Cd4 splenocytes.
|
| Data processing |
The data was RMA normalised with Affymetrix Powertools. Probes potentially
containing SNPs were excluded (25690) using the '--kill-list' tag. Annotation
files were downloaded from the Affymetrix website. The command used
was "apt-probeset-summarize
-a rma -p MoGene-1_0-st-v1.r4.pgf -c MoGene-1_0-st-v1.r4.clf -m
MoGene-1_0-st-v1.r4.mps -b MoGene-1_0-st-v1.r4.bgp --qc-probesets
MoGene-1_0-st-v1.r4.qcc --kill-list". The version of apt-probeset-summarize
used was version 1.15.1.
|
| |
|
| Submission date |
Mar 15, 2012 |
| Last update date |
Sep 23, 2019 |
| Contact name |
Hugh John French |
| E-mail(s) |
hughjohnfrench@gmail.com
|
| Organization name |
UNSW
|
| Street address |
UNSW
|
| City |
Sydney |
| State/province |
New South Wales |
| ZIP/Postal code |
2036 |
| Country |
Australia |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE36550 |
Expression data from NOD and B10 CD4 splenocytes |
|
