Profiling the transcriptomic landscape of each cell is crucial for understanding development, homeostasis, and disease. While current technologies excel in profiling gene expression in single cells, RNA products derived from these genes, known as isoforms, remain poorly understood. A single gene can produce numerous isoforms with distinct or even opposing functions, however, technological limitations have left the functional roles and biological significance largely unexplored.
To address this, we integrated long-read sequencing technologies, which are adept at profiling isoforms, with single-cell sequencing. This novel approach enables us to explore known and novel isoforms at single-cell resolution, offering insights into isoform expression across cell types and developmental stages.
We applied this method to examine isoform dynamics in the developing brain using cortical organoids and stem cell-derived cortical neurons. Our approach accurately quantified isoforms in single cells, identified distinct cell types and mapped developmental trajectories of early-born neurons. We identified 178,000 unique isoforms, including 10,000 novel ones and discovered thousands of differentially expressed genes and isoforms associated with axonogenesis and neuron development, with notable cell-type-specific isoform expression in genes such as PKM. Our Long-read scRNA-seq method is a powerful tool for profiling isoforms, defining cellular populations and elucidating complex developmental and disease processes.