Single-cell RNA sequencing of human pluripotent stem cell-derived RGC-enriched retinal cells generated from hESC and hiPSC retinal organoids

This study investigates the differentiation of retinal ganglion cells (RGCs) from human pluripotent stem cells (hPSCs) to optimise and characterise in-vitro RGC generation. Retinal organoids were derived from a human embryonic stem cell line (H9) and a laboratory-established human induced pluripotent stem cell (hiPSC) line. For each stem cell line, two independent passages of cells were generated as biological replicates (n = 2 per line). To assess transcriptional diversity and differentiation efficiency, organoids were harvested at day 40 of differentiation, dissociated into single cells, and cultured for an additional 14 days to enrich for RGCs. Following enrichment, single-cell suspensions were processed using the 10x Genomics Chromium platform and sequenced on an Illumina NovaSeq X Plus. The resulting single-cell transcriptomes provide a molecular snapshot of RGC differentiation from hPSCs and enable comparison between hESC- and hiPSC-derived retinal lineages. The data contribute to refining differentiation protocols and improving reproducibility of stem-cell-derived retinal models. Raw BCL files were processed at the Australian Genome Research Facility (AGRF) using Illumina's standard pipeline to generate FASTQ files. FASTQs were additionally processed with Cell Ranger v7.1.0 (10x Genomics) using the GRCh38/Ensembl v109 reference to produce unfiltered gene-by-cell UMI count matrices in standard 10x sparse format (barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz), which are provided as processed data in this submission. Raw FASTQ files are also archived in this ArrayExpress submission.