Spleen were mechanically dissociated and the other tissues were digested with collagenase/dispase to single cell suspensions. FoxP3+ and FoxP3- cells were sorted by flow cytometry as DAPI- TCRb+ CD4+ reporter+/- cells. Single cells were then sorted in a 96-well plate and the CEL-seq protocol was performed. mRNA was reverse transcribed using adapters containing oligo dT in each well. Single-cell cDNA libraries with different barcodes were amplified by in vitro transcription. Illumina adapters were added by reverse transcription and PCR. Library were sequenced using a HiSeq2500.
GEO Accession
GSM2949504