GSM9307805: 95P_1474; Mus musculus; RNA-Seq
Cryopreserved lung tumor cell suspensions were thawed, washed with PBS, and stained with Live/Dead Fixable Aqua Dye (Invitrogen) and surface markers in FACS Buffer. One thousand live CD45PECy7-CD8+GFP+ CAR-T cells were sorted directly into reaction buffer from the SMART-Seq mRNA Kit (Takara), and reverse transcription was performed followed by PCR amplification to generate full-length amplified cDNA. Sequencing libraries were constructed using the NexteraXT DNA sample preparation kit with unique dual indexes (Illumina) to generate Illumina-compatible barcoded libraries.  Libraries were pooled and quantified using a Qubit® Fluorometer (Life Technologies).  Sequencing of pooled libraries was carried out on a NextSeq 2000 sequencer using an XLEAP-SBS flowcell (Illumina) with paired-end 59-base reads (Illumina) with a target depth of 5 million reads per sample.