Bulk RNA-seq analysis of IFNγ-PD-L1all, IFNγ+PD-L1all and IFNγ+PD-L1hi PDAC cells

For comparison of the transcription profile between IFNγ-PD-L1all and IFNγ+PD-L1all cells, we treated KPC1199 cells with vehicle control or 10 ng/mL recombinant mouse IFNγ for 20 hours and harvested them. For comparison of PD-L1hi with PD-L1all, cells were either sorted with the top 15% PD-L1+ or total of IFNγ+PD-L1all PDAC cells, and freshly collected as soon as possible to be used for RNA preparation. For RNA preparation, PDAC cell were collected by FCM analysis of PD-L1 surface expression after the stimulation of recombinant mouse IFN-γ. RNA-seq transcriptome library was prepared following TruSeqTM RNA sample preparation Kit (Illumina) using 5 μg of total RNA, according to manufacturer’s instructions.