GSM9500046: PMP CAF culture; Homo sapiens; RNA-Seq
CAFs at passage 6 were collected by trypsinization, cryopreserved, and processed for single-cell RNA sequencing (scRNAseq). Amplified cDNA was used as input for gene expression library construction using the Chromium Next GEM Single Cell 3' v3.1 library construction reagents (10x Genomics; catalog no. 1000352). Libraries were generated through enzymatic fragmentation, end repair, A-tailing, adapter ligation, and sample index PCR amplification (10 cycles) following the manufacturer's protocol. Library quality and fragment size distribution were assessed using an Agilent Bioanalyzer, and library concentration was measured using a Qubit fluorometer. Hashtag oligonucleotide (HTO) libraries were generated from the amplified cDNA according to the 10x Genomics Feature Barcoding protocol for TotalSeq™-A antibodies. HTO fragments were selectively amplified by sample index PCR using 15 cycles. Library quality and fragment size distribution were assessed using an Agilent Bioanalyzer, and library concentrations were measured using a Qubit fluorometer.