title: GSM5112191: tumor_74_a; Mus musculus; RNA-Seq
library_construction_protocol: Samples were prepared by harvesting (1) CT2A tumors from the forebrain of mice , or (2) spleen. After single-cell preparation, cells were stained with TCR, CD4, C8, CD25, CD127. Cells were sorted on Aria 4 into 96-well plates containing RLT buffer + 1% BME. TCR+CD8+ cells were assigned to a dump channel in flow cytometry to avoid CD8 T cell contamination. TCR+CD4+CD25hi CD127lo cells were sorted as Tregs. A fraction of cells were tested for FoxP3 positivity to double-check and were all FoxP3. RNA was extracted from RLT bufffer using 2.2x SPRI bead wash. RNA was converted to cDNA,amplified, and prepared for sequencing  using the SmartSeq2 protocol (Trombetta et al., Current Protocols in Molecular Biology, 2014, 107:II:4:22).