The roundworm Caenorhabditis elegans may be best known for its cellular family tree, a series of cell divisions that produce the various cell types of the adult. The enormous effort it took in the mid-1970s to document this tree, consisting of countless hours of basic microscopy, is legendary. While methods to mark and track cells in vivo have improved over the decades, a recent set of approaches in particular has eased and extended the power of lineaging studies. They generate an extraordinary diversity of cell labels and read them out at high resolution.

Evolving barcodes offer a powerful way to resolve cell lineages.

Lineage information is important for understanding phenomena such as organogenesis, cell-fate choice, cell migration and tumor evolution. Many approaches have been developed to follow cells over time. Dyes or genetic markers can be introduced into a cell or cell population and then identified in descendants to determine lineage relationships. A second group of methods increases the number of progenitors that can be tracked; for example, sparse recombination can label cells with unique colors by producing different combinations of fluorescent reporters. Alternatively, libraries of viruses encoding a high diversity of DNA barcodes can be transfected into cells and read out in descendants by sequencing.

A new generation of tools uses the CRISPR system to evolve cellular barcodes in vivo. The Cas9 nuclease is directed by a guide RNA to specific barcode regions, where it generates a cut that can produce a unique insertion or deletion during repair; these changes then accumulate over time. Different versions use integrated GFP reporters as targets for barcoding (bioRxiv, 2017), synthetic multimerized target sites (Science 353, aaf7907-1–aaf7907-10, 2016; Nature 541, 107–111, 2017), or the guide RNA itself (Science 353, aag0511-1–aag0511-10, 2016; Nat. Methods 14, 195–200, 2017). Barcodes are read out by sequencing, or in one case, by sequential fluorescence in situ hybridization.

Variations are already being developed to provide high-resolution lineage and transcriptome readouts using single-cell RNA sequencing (bioRxiv, 2017; (bioRxiv, 2017). We look forward to further improvements in barcode fidelity and interpretation, combination with epigenetic data, control of the timing and location of barcode generation, and to challenging applications such as whole-organism lineaging.