European Research Institute for the Biology of Ageing, University of Groningen, University Medical Center Groningen, Antonius Deusinglaan 1, 9713AV, Groningen, The Netherlands.
Hubrecht Institute-KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584CT, Utrecht, The Netherlands.
Nat Commun. 2017 Dec 14;8(1):2120. doi: 10.1038/s41467-017-02214-8.
Regeneration-capable flatworms are informative research models to study the mechanisms of stem cell regulation, regeneration, and tissue patterning. However, the lack of transgenesis methods considerably hampers their wider use. Here we report development of a transgenesis method for Macrostomum lignano, a basal flatworm with excellent regeneration capacity. We demonstrate that microinjection of DNA constructs into fertilized one-cell stage eggs, followed by a low dose of irradiation, frequently results in random integration of the transgene in the genome and its stable transmission through the germline. To facilitate selection of promoter regions for transgenic reporters, we assembled and annotated the M. lignano genome, including genome-wide mapping of transcription start regions, and show its utility by generating multiple stable transgenic lines expressing fluorescent proteins under several tissue-specific promoters. The reported transgenesis method and annotated genome sequence will permit sophisticated genetic studies on stem cells and regeneration using M. lignano as a model organism.
具有再生能力的扁形动物是研究干细胞调控、再生和组织模式形成机制的有价值的研究模型。然而，缺乏转基因方法极大地限制了它们的广泛应用。在这里，我们报告了一种用于 Macrostomum lignano 的转基因方法的开发，Macrostomum lignano 是一种具有出色再生能力的基础扁形动物。我们证明，将 DNA 构建体微注射到受精卵的单细胞阶段，然后进行低剂量辐射，通常会导致转基因随机整合到基因组中，并通过生殖系稳定传递。为了方便选择转基因报告基因的启动子区域，我们组装并注释了 M. lignano 基因组，包括转录起始区域的全基因组图谱，并通过生成多个稳定的表达荧光蛋白的转基因系，展示了其在几种组织特异性启动子下的用途。报道的转基因方法和注释的基因组序列将允许使用 M. lignano 作为模式生物进行干细胞和再生的复杂遗传研究。
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