GSM8229996: Sort-purified (CD8/CD45.1) TCR OT1 CD8 T cells from B16-OVA-GP61 tumors (no TCR SMARTA CD4 T cells) replicate#1; Mus musculus; RNA-Seq
CD8 T cells were FACS-sorted straight into Trizol tube and stored at -80°C until all samples were collected. RNA from sorted cells was extracted using RNeasy mini kit (Qiagen) according to instructions provided by the manufacturer. After ribogreen quantification and quality control of Agilent BioAnalyzer, 6-15 ng of total RNA underwent amplification (12 cycles) using the SMART-seq V4 (Clonetech) ultralow input RNA kit for sequencing. 10ng of amplified cDNA was used to prepare Illumina hiseq libraries with the Kapa DNA library preparation chemistry (Kapa Biosystems) using 8 cycles of PCR. Samples were barcoded and run on a Hiseq 2500 1T in a 50bp/50bp Paired end run, using the TruSeq SBS Kit v3 (Illumina).  An average of 51 million paired reads were generated per sample and the percent of mRNA bases was 62.5% on average. 10ng of amplified cDNA was used to prepare Illumina hiseq libraries with the Kapa DNA library preparation chemistry (Kapa Biosystems) using 8 cycles of PCR. Samples were barcoded and run on a Hiseq 2500 1T in a 50bp/50bp Paired end run, using the TruSeq SBS Kit v3 (Illumina).