title: GSM5100141: P81p4_Ebo; Homo sapiens; RNA-Seq
library_construction_protocol: For isolation of Ebo, Em and Eff CD8 T cell subsets, 1-2 106 tumor-infiltrating leukocytes were resuspended in 200 ml staining buffer containing fluorophore-conjugated mAb cocktail against human cell surface molecules: CD45 BV510 (clone HI30), CD8 PerCP-Cy5.5 (clone RPA-T8) , CCR7 BV421 (clone G043H7), CD45RO PE (cloneUCHL1) and CD45RA PE-Cy7 (clone HI100). After 20 min incubation at 4 °C, cells were washed twice, and cell sorting was performed using a BD FACS Aria III. Approximately 1 × 104, 1.5-2× 104 and 0.5-1 × 104 of Ebo, Em and Eff cells were sorted respectively as an average across experiments. In three experiments cells were directly sorted in lysis buffer RLT (QIAGEN, RNeasy® Micro Kit) which already added b-ME and DTT, and the lysis cells were extracted total RNA follow the RNeasy® Micro Kit (QIAGEN, cat. 74004) protocol. the total RNA samples were submit to Yale Center for Genome Analysis (YCGA) for analyzed QC  and used for RNA sequencing.