BASELINE: A CRISPR Base Editing Platform for Mammalian-Scale Single-Cell Lineage Tracing

A cells 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 to map the importance of lineage, yet 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 within 50 synthetic target sites, which are integrated multiple times into a cells genome. We show 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. Single-cell sequencing reveals high-fidelity capture of these lineage recorders, recovering lineage reconstructions up to 46 cell-divisions deep, within the estimated range of mammalian development. We expect BASELINE to be applicable to various lineage-tracing projects in development and disease, especially in which cellular engineering makes small, more distributed systems challenging.