Unraveling the fates of resident stem cells during tissue regeneration is an important objective in clinical and basic research. Genetic lineage tracing based on Cre-loxP recombination provides an effective strategy for inferring cell fate and cell conversion in vivo. However, the determination of the exact fates of resident stem cells or their derivatives in disease states and during tissue regeneration remains controversial in many fields of study, partly because of technical limitations associated with Cre-based lineage tracing, such as, for example, off-target labeling. Recently, we generated a new lineage-tracing platform we named DeaLT (dual-recombinase-activated lineage tracing) that uses the Dre-rox recombination system to enhance the precision of Cre-mediated lineage tracing. Here, we describe as an example a detailed protocol using DeaLT to trace the fate of c-Kit⁺ cardiac stem cells and their derivatives, in the absence of any interference from nontarget cells such as cardiomyocytes, during organ homeostasis and after tissue injury. This lineage-tracing protocol can also be used to delineate the fate of resident stem cells of other organ systems, and takes ~10 months to complete, from mouse crossing to final tissue analysis.

中文翻译：

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DeaLT对驻留干细胞的遗传谱系追踪。

揭示组织再生过程中常驻干细胞的命运是临床和基础研究的重要目标。基于Cre-loxP重组的遗传谱系追踪为推断体内细胞命运和细胞转化提供了有效的策略。但是，在许多研究领域中，确定疾病状态和组织再生过程中驻留干细胞或其衍生物的确切命运仍然存在争议，部分原因是基于Cre的谱系追踪技术存在局限性，例如， -目标标签。最近，我们生成了一个名为DeaLT（双重重组酶激活的谱系追踪）的新谱系追踪平台，该平台使用Dre-rox重组系统来增强Cre介导的谱系追踪的精度。这里，⁺心脏干细胞和它们的衍生物，在器官内稳态中没有来自非靶细胞，例如心肌细胞，任何干扰的和组织损伤之后。此谱系追踪方案也可用于描述其他器官系统中常驻干细胞的命运，从小鼠杂交到最终组织分析需要约10个月的时间才能完成。