BASELINE: a CRISPR base editing platform for mammalian-scale single-cell lineage tracing

A cell's fate is shaped by its inherited state, or lineage, and the ever-shifting context of its environment. CRISPR-based recording technologies are a promising solution for mapping the lineage of a developing system; however, challenges remain regarding single-cell recovery, engineering complexity, and scale. Here, we introduce BASELINE, which uses base editing to generate high-resolution lineage trees in conjunction with single-cell profiling. BASELINE uses the Cas12a adenine base editor to irreversibly edit nucleotides across target arrays built from 50 synthetic target sites, which are integrated multiple times into a cell's genome. We demonstrate that BASELINE accumulates lineage-specific marks over a wide range of biologically relevant intervals, recording more than 4300 bits of information in a model of pancreatic cancer, a 50-fold increase over existing technologies. Single-cell sequencing reveals high-fidelity capture of these recorders, averaging 29 cell divisions captured per lineage, within the estimated range of mammalian development. We expect BASELINE to apply to a wide range of lineage-tracing projects in development and disease, especially those in which cellular engineering makes small, more distributed systems challenging.