MC38 tumor sample treated with Alistipes.fingegodii for single cell TCRseq (CRX1824297; CRA027345)
To perform scTCR-seq on fresh tumor tissues, single-cell suspension was prepared as mentioned above. Single-cell suspension was then loaded onto a droplet-based platform (10X Genomics) according to the manufacturer's instructions. cDNA libraries were prepared using the Chromium Single Cell 5' v2 Reagent Kit and Gel Bead Kit, Single Cell V(D)J Enrichment Kit, Human T Cell. Briefly, single cells were suspended in PBS containing 0.04% BSA. Captured cells were lysed, and the released RNA was barcoded via reverse transcription within individual Gel Bead-In Emulsions (GEMs). Reverse transcription was performed on a S1000 Touch Thermal Cycler (Bio-Rad), followed by paired-end 150 bp sequencing on an Illumina NovaSeq platform at a depth of at least 100,000 reads per cell.