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Sample GSM4273430 Query DataSets for GSM4273430
Status Public on Dec 18, 2020
Title d30 off 2
Sample type RNA
Source name CD4+ T cells primed in vivo, then exposed to different levels of persisting antigen
Organism Mus musculus
Characteristics cells: sorted CD4+ AND T cells
days after transfer: 30
doxycycline [ug/ml] day -1 to 3: 100
doxycycline [ug/ml] day 3 to 10 or 30: day 3 to 10: 100 day 10 to 30: 0
Treatment protocol The recipients had been treated one day before the transfer with an anti-CD40 antibody to activate dendritic cells and with 100 µg/ml doycycline in the drinking water which induces the expression of the cDNA construct in dendritic cells. One group (none) was then withdrawn from doxycycline. We have seen before that the T cells in recipients undergoing this treatment will develop into Th1 memory cells (Obst et al., 2007). Another group (low) was switched to 10 µg/ml while a third (intermediate) was maintained on the 100 µg/ml dose. In addition, another strain of mice (cMCC) were used as recipients that expressed an invariant chain/MCC construct that is constitutively expressed in MHC-II+ cells at high levels (high). The adoptively transferred AND TCR tg T cells were sorted twice on day 10 as CD4+CD90.2+CD45.1+ cells directly into Trizol LS , mixed and stored at -80°C for processing. - A second series of sorts was done 30 days after transfer. Instead of the cMCC recipients, in which the AND T cells undergo apoptosis by then, iMCC animals were used which were taken off doxycycline on day 10 (off). Naive control AND T cells were transferred 3 days before the sort into non-transgenic recipients.
Growth protocol AND TCR transgenic CD45.1+ Rag2o/o CD4+ T cells were adoptively transferred into iMCC recipients carrying two transgenes that allow for the dynamic regulation of the expression of an invariant chain cDNA which carries a moth cytochrome c (MCC) epitope instead of the CLIP sequence.
Extracted molecule total RNA
Extraction protocol The Trizol samples were thawed on ice, 300 µl chloroforn was added, shaken, incubated for 3 minutes at room temperature and spun at 12,000 x g and 4°C. The aqueous phase was transferred to new tubes, 1 volume of isopropanol and 1 µl of linear polyachrylamide stock solution (25 µg/µl) was added. The samples were precipitated for 16 hours at -20°C and centrifuged for 10 minutes at 12,000 x g and 4°C. Supernatants were removed and RNA pellets washed with 70% ethanol. RNA pellets were resuspended in 100 µl RNAse-free water and purified with NucleoSpin RNA Clean-up XS columns as per manufacturer's instructions, from which the RNA was eluted with 12 µl water and stored at -80°C. RNAs were quantified with an Agilent 2100 Bioanalyzer. All samples had RNA integrity values > 8 and were use for amplification and microarray hybridizations.
Label biotin
Label protocol Total RNA (0.1-30 ng) was amplified and labeled using the Ovation Pico WTA System V2 in combination with the Encore Biotin Module (Nugen).
Hybridization protocol Amplified cDNA was hybridized on Affymetrix Mouse Gene ST 2.0 arrays containing about 35,000 probe sets.
Scan protocol Staining and scanning using a GeneChip Scanner 3000 7G was done according to the Affymetrix expression protocol including minor modifications as suggested in the Encore Biotin protocol.
Data processing R/Bioconductor, rma normalization, default parameters
Submission date Jan 15, 2020
Last update date Dec 18, 2020
Contact name Tobias Straub
Organization name LMU Munich
Department Biomedical Center, Bioinformatics
Street address Großhadener Str. 9
City Martinsried
ZIP/Postal code 82152
Country Germany
Platform ID GPL16570
Series (2)
GSE143733 Gene expression of CD4+ T cells in response to antigen persisting at different doses for different times
GSE143739 Gene expression of CD4+ T cells

Data table header descriptions
VALUE RMA signal intensity

Data table
17200001 3.59
17200003 4.03
17200005 2.35
17200007 3.11
17200009 4.42
17200011 3.17
17200013 2.56
17200015 3.4
17200017 3.62
17200019 2.58
17200021 2.54
17200023 3.85
17200025 4.26
17200027 5.05
17200029 2.98
17200031 2.6
17200033 3.71
17200035 2.86
17200037 2.97
17200039 3.16

Total number of rows: 41345

Table truncated, full table size 560 Kbytes.

Supplementary file Size Download File type/resource
GSM4273430_d30_off_2.CEL.gz 8.3 Mb (ftp)(http) CEL
Processed data included within Sample table

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