single nucleus RNA sequencing for donor 3848; library 1
Droplet-based libraries were generated using the Next Gem Single Cell 3' v3 or v3.1 reagent kits (10x Genomics) and chromium controller according to manufacturer's instructions. Resulting libraries were indexed with the KAPA Unique Dual-Indexed Adapter Kit (Roche KK8726) and sequenced on an Illumina Novaseq 6000 with 150 paired-end reads by Genuity Sciences (Dublin, Ireland). Samples were prepared in batches of up to six donors at a time that always included male and female donors as well as mixed ages (see Table S3). To prevent age or gender bias in our batches, some samples have multiple biological replicates, prepared on different dates. A single replicate each from three distinct donors clustered abnormally during downstream analysis and was therefore excluded from analysis. After filtering, the only clusters exhibiting batch bias are those which are infant-specific and biologically driven (Figure S1). Because those cells were only present in infant donors, the only batches contributing to those clusters are those which included an infant.