RNA-sequencing of three murine memory CD8 T cell subsets: IL-7Rα+KLRG1- , IL-7Rα+KLRG1+, and IL-7Rα-KLRG1+

The goal of this study is to characterize the transcriptional profile of three memory CD8 T cell subsets: IL-7Rα+KLRG1- (memory precursors, MP), IL-7Rα+KLRG1+ (KLRG1+ MP) and IL-7Rα-KLRG1+ (terminal effectors, TE). Splenocytes were isolated from mice that had been infected with LCMV Armstrong 50 days prior (2×105 pfu, intraperitoneally). Cells were stained with extracellular antibodies (CD4, CD8, CD19, IL-7Rα, and KLRG1 from Biolegend and SouthernBiotech) and an LCMV-GP33 tetramer for 30 minutes. Cell viability was determined with the Live/Dead fixable aqua dead cell stain kit (Invitrogen). Cells were washed with PBS + 2% FBS and sorted on a FACSAria II (BD Biosciences). Three populations of GP33+ CD8 T cells were sorted in triplicate: IL-7Rα+KLRG1- (MP), IL-7Rα+KLRG1+ (KLRG1+ MP), and IL-7Rα-KLRG1+ (TE). RNA was isolated from each sample using the All-Prep DNA/RNA Micro Kit (Qiagen) and then submitted for sequencing.