LoxCode in vivo barcoding reveals epiblast clonal fate bias to fetal organs

Much remains to be learned about the clonal fate of mammalian epiblast cells. Here, we develop high-diversity Cre recombinase-driven LoxCode barcoding for in vivo clonal lineage tracing for bulk tissue and single-cell readout. Embryonic day (E) 5.5 pre-gastrulation embryos were barcoded in utero, and epiblast clones were assessed for their contribution to a wide range of tissues in E12.5 embryos. Some epiblast clones contributed broadly across germ layers, while many were biased toward either blood, ectoderm, mesenchyme, or limbs, across tissue compartments and body axes. Using a stochastic agent-based model of embryogenesis and LoxCode barcoding, we inferred and experimentally validated cell fate biases across tissues in line with shared and segregating differentiation trajectories. Single-cell readout revealed numerous instances of asymmetry in epiblast contribution, including left-versus-right and kidney-versus-gonad fate. LoxCode barcoding enables clonal fate analysis for the study of development and broader questions of clonality in murine biology.