Single-Molecule Barcoding Technology for Single-Cell Genomics

Recent advances in barcoding technologies have introduced unprecedented scalability to single-cell genomic experiments. However, most large-scale studies are focused on the generation of reference cell atlases, which prioritize this goal over cost-efficiency and logistics. For large-scale, multi-million-cell studies to be routine in fields such as clinical diagnostics, drug/target discovery, genetic screening, and gene therapy, it is crucial to simplify the single-cell workflow and the generation of barcoding reagents. Here, we introduce AmpliDrop, a technology that streamlines single-cell barcoding by initiating the process using inexpensive single-copy barcodes, which are then amplified as part of the library preparation process, rather than relying on pre-synthesized multi-copy barcode pools. This strategy eliminates the need for barcoded beads and complex combinatorial indexing workflows. Additionally, cell compartmentalization is achieved using a conventional electronic pipette or a robotic liquid handler for automation, providing flexibility for any scale and number of samples. AmpliDrop can capture transcriptomes and chromatin accessibility and can also accommodate user-customized modalities for protein profiling, bacterial or viral DNA detection, and genetic perturbations without construct limitations. We showcase the discovery potential of AmpliDrop by investigating the influence of physical perturbation on cell diversity in human cortical organoids, revealing the selective vulnerability of glycolytic radial glia.