title: Illumina HiSeq 1500 sequencing; RNA-seq of murine CD4 T cells after pentanoate treatment compared to untreated control
design: RNA-seq of murine CD4 T cells after pentanoate treatment compared to untreated control
library_name: CD4_T_cells_control-2_s
library_construction_protocol: Spleen and lymphnodes were isolated from FIR/ TIGER mice on C57/Bl6 Background (IL-10/ GFP - Foxp3/ RFP -reporter mouse). Cells were homogenized with 30 M cell strainers, CD4 T cells were isolated using Magnet-associated cell sorting (negative selection) to a purity of over 90% A 48-well plate was coated with Anti-CD3-Ab (5 g/ml) overnight the day before; cells were cultured in RPMI complete (500000 cells/ well in 500 L) conditioned with plate-bound Anti-CD3-antibody, IL-2, IL-23, IL-1beta, Anti-IL-4-Antibody, Anti-IFN-gamma-Antibody and Anti-CD28-Antibody in presence or absence (control) of 4 mM pentanoate (valeric acid) for 3 days at 37C, 5% CO2 saturation and 95% humidity. Cells were harvested on day 3 and RNA was isolated  Cells were harvested and washed in PBS, RNA-isolation was performed according to the protocol of the Qiagen RNeasy Mini Kit (Qiagen) incl. DNase digestion RNASeq libraries were prepared using a TruSeq Stranded mRNA Sample Prep kit (Illumina, Tokyo, Japan).
attributes.tag: Experimental Factor: compound
attributes.value: none
attributes.tag: Experimental Factor: dose
attributes.value: 0