Chromium single cell 3 library gel bead kit v2 or v3

Single-cell RNAseq libraries were prepared using a Chromium Single Cell 3′ Library & Gel Bead Kit v2 or v3 (10X Genomics), according to the manufacturer’s protocol (user guide). Two chips were loaded with the accurate volumes calculated based on the 'Cell Suspension Volume Calculator Table.' The initial single-cell suspension being estimated at >600 cells/μl, we targeted to recover a maximum 10,000 cells. Once GEMs were obtained, reverse transcription and cDNA amplification steps were performed. Sequencing was done on Illumina NovaSeq 6000 S2 flow cell generating paired-end reads. Different sequencing cycles were performed for the different reads, R1 and R2. R1 contained 10× barcodes and UMIs, in addition to an Illumina i7 index, and R2 contained the transcript-specific sequences. All steps were performed at the Next Generation Sequencing platform at the University of Bern.