B7S1 negatively regulates T cells and its expression correlates with poor prognosis of cancer patients. In order to understand how B7S1 signaling contributes to dysfunction of CD8+ T cell in the TME, we conducted transcriptional analysis of OVA-specific CD8+ TILs and different TIL subsets from E.G7-bearing WT and B7S1 KO mice (Day 21). Overall design: One million E.G7 were inoculated subcutaneously into mice on Day 0. On Day 21, OVA-specific CD8+ TILs from E.G7-bearing WT and B7S1 KO mice (6 mice per group), or PD-1+Tim-3-B7S1R+(B7S1R_SP), PD-1+Tim-3+B7S1R-(TIM-3_SP), PD-1+Tim-3-B7S1R-(DN) CD8+ TILs from E.G7-bearing WT or B7S1 KO mice were sorted into lysis buffer using BD FACSAria Cell Sorter. cDNA was made and library was constructed using SMARTer Ultra Low Input RNA for Illumina Sequencing-HV kit and Creator SMART cDNA Library Construction Kit (Clontech) according to manufacturer’s instructions. The library products were sequenced via Illumina HiSeq 2500/4000 and successfully mapped to mouse genome. Reads counts were normalized based on RPKM/FPKM, fold changes were calculated for all possible comparisons.