MOLECULAR BASIS OF PLANARIAN REGENERATION

这项提议的目标是使用后生动物作为后生动物模型 解剖和操纵动物分子基础的系统 再生。行星动物被选为模型系统是因为 它们在后生动物系统发育中的关键地位，它们显著的再生 能力，以及这些动物再生的全潜能性质 干细胞群体(新生母细胞)。首先，我们建议确定和 描述与指定和维持前房相关的基因 和后天再生的命运。我们将实现这一目标 使用基于聚合酶链式反应的基因筛查。方法论 涉及再生和再生的cDNA的消减杂交 不能再生的组织，然后扩增出独特的富含的cDNA 到每个组织(见初步结果)。基因筛选分离 CDNA的特征是测序、发育的北方和 原位杂交。其次，我们将利用无性恋者 和有性繁殖策略，以及它们的 全能再生干细胞以开发方法学 是生产转基因动物所必需的。转基因将 允许对其序列、发育中的基因进行功能测试 表达谱和组织特异性表明潜在的调控 在再生过程中扮演的角色。一旦确定， 浮游生物再生特异性基因的功能特征 将垂直整合到高等再生的研究中 有机体。最终，在行星动物身上发现的基因，以及他们的 再生过程中的交互作用将定义一系列有用的 有助于解开更复杂的表观遗传学的分子模板 脊椎动物再生的过程，并揭示导致 在一些后生动物中允许再生，但不是所有后生动物。

The objective of this proposal is to use planarians as a metazoan model system in which to dissect and manipulate the molecular basis of animal regeneration. Planarians have been chosen as a model system because of their key position in metazoan phylogeny, their remarkable regeneration abilities, and the totipotential nature of these animals' regenerative stem cell population (neoblasts). First, we propose to identify and characterize the genes involved in specifying and maintaining anterior and posterior regeneration fates. We will accomplish this objective using a polymerase chain reaction-based gene-screen. The methodology involves the subtractive hybridization of cDNAs from regenerating and non-regenerating tissues, and then amplifying the enriched cDNAs unique to each tissue (see preliminary results). The gene-screen-isolated cDNAs will be characterized by sequencing, developmental Northerns and in situ hybridizations. Secondly, we will take advantage of the asexual and sexual reproductive strategies of planarians, and of their totipotential regenerative stem cells to develop the methodologies necessary for the production of transgenic animals. Transgenesis will allow functional testing of those genes whose sequence, developmental expression profile, and tissue specificity indicate potential regulatory roles during the process of regeneration. Once identified, the functional characterization of regeneration-specific genes in planarians will be vertically integrated to the study of regeneration in higher organisms. Ultimately, the genes identified in planarians, and their interactions during regeneration will define a series of useful molecular templates which will help unravel the more complex epigenetic processes of vertebrate regeneration, and uncover the factors which make regeneration permissive in some, but not all metazoans.