GSM6995624: colonTreg_3-3_GvHD_Tcra_R1; Mus musculus; OTHER
Total cellular RNA was isolated from Treg (freshly isolated, in vitro expanded, or re-isolated after BMT) using the RNeasy Mini Kit or the RNeasy Micro Kit (Qiagen). Following the manufacturer's recommendations, samples originating from 500 to 500k cells were processed with the Micro Kit whereas the Mini Kit was applied to all samples exceeding 500k cells. RNA concentration and quality were simultaneously assessed using RNA Screentape Analysis or High Sensitivity RNA Screentape Analysis (Agilent), depending on the expected yield. Total cellular RNA was obtained as described in section “RNA-seq library preparation”. TCR repertoire sequencing libraries for Illumina sequencing were prepared on the basis of {Mamedov, 2013 #7} and the protocol was adapted, as follows. For the generation of 5'RACE-ready cDNA total RNA ranging from 10 - 200 ng (low input) or 200 - 1000 ng (high input) in a total of 10 µl RNAse-free water was combined with 1 µ