Stable genomic integration of exogenous transgenes is essential in neurodevelopmental and stem cell studies. Despite tools driving increasingly efficient genomic insertion with DNA vectors, transgenesis remains fundamentally hindered by the impossibility of distinguishing integrated from episomal transgenes. Here, we introduce an integration-coupled On genetic switch, iOn, which triggers gene expression upon incorporation into the host genome through transposition, thus enabling rapid and accurate identification of integration events following transfection with naked plasmids. In vitro, iOn permits rapid drug-free stable transgenesis of mouse and human pluripotent stem cells with multiple vectors. In vivo, we demonstrate faithful cell lineage tracing, assessment of regulatory elements, and mosaic analysis of gene function in somatic transgenesis experiments that reveal neural progenitor potentialities and interaction. These results establish iOn as a universally applicable strategy to accelerate and simplify genetic engineering in cultured systems and model organisms by conditioning transgene activation to genomic integration.

外源转基因的稳定基因组整合在神经发育和干细胞研究中至关重要。尽管利用DNA载体驱动工具越来越有效地进行基因组插入，但转基因仍从根本上受到阻碍，无法区分整合型和游离型转基因。在这里，我们介绍了一个整合耦合的On遗传开关iOn，它通过转座整合到宿主基因组中时触发​​基因表达，从而能够在裸质粒转染后快速，准确地识别整合事件。在体外，iOn允许使用多种载体对小鼠和人多能干细胞进行快速无药物稳定的转基因。体内，我们在揭示神经祖细胞潜力和相互作用的体细胞转基因实验中展示了忠实的细胞谱系追踪，调节元件评估以及基因功能的镶嵌分析。这些结果将iOn确立为一种普遍适用的策略，可通过调节转基因激活到基因组整合来加速和简化培养系统和模型生物中的基因工程。