title: GSM7675404: liver,replicate4; Mus musculus; RNA-Seq
library_construction_protocol: Livers were chopped and incubated in RPMI + 10% FBS with collagenase VIII (100U/mL; Sigma Aldrich)and DNase I (150 µg/mL; Sigma Aldrich) for 45 minutes at 37° C and then passed through a 70-µm cell strainer (Fisher Scientific). Leukocytes were enriched for by density-gradient centrifugation over a 40/80% Percoll (GE Healthcare) gradient, and remaining red blood cells were lysed with ACK lysis buffer (Lonza) and washed in PBS. Spleens were removed and placed in RPMI + 10%FBS. Spleens were then passed through a 70-µm cell strainer (Fisher Scientific). Red blood cells were lysed with ACK lysis buffer (Lonza) and washed in PBS. For both tissues, cells were sorted on singlet cells, live cells, CD45+ cells, DumpNEG (CD11b+, CD11c+, CD49b+, CD19+, CD8+) cells, CD3+ cells, CD4+ cells. Cells were isolated from 2 mice and pooled together, with equal numbers of cells pooled from each animal used. 12,000 cells from each tissue were loaded on a 10X Genomics Next GEM chip and single-cell GEMs were generated on a 10X Chromium Controller. Subsequent steps to generate cDNA and sequencing libraries were performed following 10X Genomics' protocol. Libraries were pooled and sequenced using Illumina NovaSeq SP 100 cycle as per 10X sequencing  recommendations.