title: GSM7207907: TIL_Prdm1-/-_UT_rep2; Mus musculus; RNA-Seq
library_name: GSM7207907
library_construction_protocol: Following dissection, tumors were mechanically disrupted using scissors, followed by digestion with 0.33 mg/ml Liberase TL (Roche) and 0.2 mg/ml DNase (Roche) in serum-free RPMI at 37°C for 30 min. Tumors were dispersed through a 70 μm filter using PBS + 2 mM EDTA and cells resuspended in cRPMI. Lymphocytes were enriched from tumor samples by centrifugation using a Histopaque 1119 density gradient.  (Sigma-Aldrich). Cells were stained using directly conjugated antibodies against extracellular antigens and Live, CD3+ cells sorted. Single cell RNA sequencing was carried out using the using the Chromium Single Cell 5' V(D)J Reagent kit, v1 and v2 (10X Genomics). Purified single cell suspensions were loaded onto the 10X Chromium Controller and libraries prepared as per the manufacturer's instructions, using AMPure beads for size selection and purification and the Bioanalyzer High sensitivity DNA assay (Agilent) to assess cDNA concentration and size of final libraries. Library concentrations were quantified by qPCR using the Illumina Library Quantification kit (KAPA). Libraries were pooled for sequencing at a final concentration of 10 nM. Pooled libraries were sequenced using a NextSeq High output run or on the NovaSeq platform using 26 x 91 paired end reads.