Thymus from each mouse was dissected and homogenized into single-cell suspensions for cell sorting. Treg (CD4+CD8-CD25+GFP+) and non-Treg (CD4+CD8-GFP-) cells were sorted by a MoFlo-XDP (Beckman Coulter) and total RNA was extracted with RiboEx (GeneAll, Seoul, Republic of Korea) according to the manufacturer's protocol. Reverse transcription and subsequent PCR amplification (RT-PCR1) is performed using Qiagen OneStep RT PCR mix (Qiagen). The cDNA is selected and unused primer is removed by SPRIselect bead selection (Beckman Coulter) followed by a second round of amplification performed with a pair of primers that are specific for communal sites engineered onto the 5' end of the C- and V- primers used during RT-PCR1. The final constructed library includes Illumina sequencing adapters and a 6-nucleotide internal barcode associated with the C-gene primer.