Morris Joshua, Ladurner Peter, Rieger Reinhard, Pfister Daniela, Del Mar De Miguel-Bonet Maria, Jacobs David, Hartenstein Volker
Department of Molecular, Cell and Developmental Biology, University of California Los Angeles, Los Angeles, CA 90095, USA.
Dev Genes Evol. 2006 Nov;216(11):695-707. doi: 10.1007/s00427-006-0098-z. Epub 2006 Oct 5.
We report the development of an Expressed Sequence Tag (EST) resource for the flatworm Macrostomum lignano. This taxon is of interest due to its basal placement within the flatworms. As such, it provides a useful comparative model for understanding the development of neural and sensory organization. It was anticipated on the basis of previous studies [e.g., Sánchez-Alvarado et al., Development, 129:5659-5665, (2002)] that a wide range of developmental markers would be expressed in later-stage macrostomids, and this proved to be the case, permitting recovery of a range of gene sequences important in development. To this end, an adult Macrostomum cDNA library was generated and 7,680 Macrostomum ESTs were sequenced from the 5' end. In addition, 1,536 of these aforementioned sequences were sequenced from the 3' end. Of the roughly 5,416 non-redundant sequences identified, 68% are similar to previously reported genes of known function. In addition, nearly 100 specific clones were obtained with potential neural and sensory function. From these data, an annotated searchable database of the Macrostomum EST collection has been made available on the web. A major objective was to obtain genes that would allow reconstruction of embryogenesis, and in particular neurogenesis, in a basal platyhelminth. The sequences recovered will serve as probes with which the origin and morphogenesis of lineages and tissues can be followed. To this end, we demonstrate a protocol for combined immunohistochemistry and in situ hybridization labeling in juvenile Macrostomum, employing homologs of lin11/lim1 and six3/optix. Expression of these genes is shown in the context of the neuropile/muscle system.
我们报告了扁形虫利氏大口涡虫（Macrostomum lignano）表达序列标签（EST）资源的开发情况。由于该分类单元在扁形虫中处于基础地位，因此备受关注。如此一来，它为理解神经和感觉组织的发育提供了一个有用的比较模型。基于先前的研究[例如，桑切斯 - 阿尔瓦拉多等人，《发育》，129:5659 - 5665，（2002）]预计，一系列发育标记物会在后期的大口涡虫中表达，事实证明确实如此，从而得以获取一系列对发育重要的基因序列。为此，构建了一个成年利氏大口涡虫的cDNA文库，并从5'端对7680个利氏大口涡虫EST进行了测序。此外，从3'端对上述序列中的1536个进行了测序。在鉴定出的约5416个非冗余序列中，68%与先前报道的已知功能基因相似。此外，还获得了近100个具有潜在神经和感觉功能的特定克隆。基于这些数据，一个注释可搜索的利氏大口涡虫EST集合数据库已在网上提供。一个主要目标是获得能够重建基础扁形虫胚胎发生，特别是神经发生的基因。回收的序列将作为探针，借此追踪谱系和组织的起源及形态发生。为此，我们展示了一种在幼年利氏大口涡虫中进行免疫组织化学和原位杂交联合标记的方案，使用了lin11/lim1和six3/optix的同源物。这些基因的表达在神经纤维网/肌肉系统的背景下得以展示。
