Sydney Hart

Functionally characterizing disease associated genetic variants in relevant cellular contexts remains a critical bottleneck in translating genomic associations into therapeutic opportunity. To address this, we developed a custom targeted DNA and RNA assay using the Tapestri® Single-Cell Platform, enabling simultaneous capture of CRISPR-edited genotypes and downstream transcriptional responses in primary human immune cells. Using isogenic CRISPR-edited cells, this approach isolates the effect of a single nucleotide change from background genomic variation, providing a more direct link between variant and biological mechanism than population-level association studies alone. We benchmarked three CRISPR editing strategies — prime editing, base editing, and HDR — across disease associated loci in PBMCs and T-cells, demonstrating editor-specific differences in on-target precision, off-target activity, and downstream transcriptional impact. Using this platform, we show that single-nucleotide changes at autoimmune-implicated loci produce disease-relevant, stimulation-dependent transcriptional consequences in primary immune cells, establishing a proof-of-concept for functionally connecting genetic variation to disease mechanism at single-cell resolution