High-resolution maps of embryonic development suggest that acquisition of cell identity is not limited to canonical germ layers but proceeds via alternative routes. Despite evidence that visceral organs are formed via embryonic and extra-embryonic trajectories, the production of organ-specific cell types in vitro focuses on the embryonic one. Here we resolve these differentiation routes using massively parallel single-cell RNA sequencing to generate datasets from FOXA2^Venus reporter mouse embryos and embryonic stem cell differentiation towards endoderm. To relate cell types in these datasets, we develop a single-parameter computational approach and identify an intermediate en route from extra-embryonic identity to embryonic endoderm, which we localize spatially in embryos at embryonic day 7.5. While there is little evidence for this cell type in embryonic stem cell differentiation, by following the extra-embryonic trajectory starting with naïve extra-embryonic endoderm stem cells we can generate embryonic gut spheroids. Exploiting developmental plasticity therefore offers alternatives to pluripotent cells and opens alternative avenues for in vitro differentiation.

胚胎发育的高分辨率图表明，细胞身份的获得并不局限于典型的胚层，而是通过替代途径进行的。尽管有证据表明内脏器官是通过胚胎和胚胎外轨迹形成的，但体外器官特异性细胞类型的产生主要集中在胚胎细胞上。在这里，我们使用大规模并行单细胞 RNA 测序从 FOXA2 ^{Venus生成数据集来解析这些分化途径}报告小鼠胚胎和胚胎干细胞向内胚层的分化。为了关联这些数据集中的细胞类型，我们开发了一种单参数计算方法，并确定了从胚胎外身份到胚胎内胚层的中间过程，我们在胚胎第 7.5 天在胚胎中进行了空间定位。虽然在胚胎干细胞分化中几乎没有这种细胞类型的证据，但通过遵循从幼稚的胚胎外内胚层干细胞开始的胚胎外轨迹，我们可以生成胚胎肠道球体。因此，利用发育可塑性提供了多能细胞的替代品，并为体外分化开辟了替代途径。