I7 index

scRNA-seq libraries were prepared using the Chromium Single Cell 3’ Library and Gel Bead Kit v3 (10X Genomics), according to the manufacturer’s protocol (User Guide). Chips were loaded after calculating the accurate volumes using the ‘Cell Suspension Volume Calculator Table’. With an initial single-cell suspension concentration equal to 1000 cells/µl, we targeted to recover approximately 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 10X barcodes and UMIs, in addition to an Illumina i7 index. While R2 contained the transcript-specific sequences.

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.

Libraries for scRNA-seq were prepared using the Chromium Single Cell 3′ Library and Gel Bead Kit v3 (10X Genomics), according to the manufacturer’s protocol (User Guide). Briefly, wells were loaded after calculating single-cell suspension concentration of each sample equal to 1,200 cells/μl. Targeted recovery rate was approximately 10,000 cells. 10X Chromium Chip performed GEMs generation, reverse transcription, and cDNA amplification. Illumina NovaSeq 6000 S2 flow cell was used for deep sequencing generating paired-end reads. Different sequencing cycles were performed for the different reads, R1 and R2. R1, contained 10X barcodes and UMIs, in addition to an Illumina i7 index. R2, contained the transcript-specific sequences.