RPG: The Isolation and Characterization of Recombination Activating Genes (RAGs) from a Lower Chordate

RPG：下脊索动物重组激活基因 (RAG) 的分离和表征

• 批准号: 9629300
• 负责人: Simona Bartl
• 金额: $ 2.29万
• 依托单位: University of North Carolina at Wilmington
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-08-01 至 1998-10-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9629300&HistoricalAwards=false
• 关键词: RPG; Isolation; Characterization; Recombination; Activating

Bartl MCB-9629300 Recombination activating genes (RAG-1 and RAG-2) are essential for gene rearrangement in mammals. Previously isolated PCR products from nurse shark DNA that have sequence similarity to mammalian RAG-1 genes will be used to screen a genomic library. Since mammalian, avian and amphibian RAG-1 and RAG-2 are tightly linked, the same may be true of the shark homologs. Genomic clones containing shark RAG-1 will be assayed for linked RAG-2 homologs using PCR, heterologous probes or by searching for an open-reading frame. Following the proposed planning period, the shark RAG-1 and RAG-2 homologs will be used to assay expression in lymphoid organs from nurse sharks of different ages using Northern blots and in situ hybridization. Recombination activity of shark RAG proteins will be assessed in mammalian fibroblasts using artificial substrates. The immune system of higher vertebrates is a complex network of cells, organs and tissues. The intricate workings and the evolution of such a tightly linked system are not well understood. A basic process required for immune function is the rearrangement of subgenic elements to form functional antigen receptors. Elucidating the mechanism in organisms distantly related to mammals is crucial for our understanding of the evolution of adaptive immunity. For the proposed research plan, shark is selected because they are the taxon most distantly related to mammals in which an adaptive immune system has been documented.

Bartl MCB-9629300 重组激活基因（RAG-1 和 RAG-2）对于哺乳动物的基因重排至关重要。 先前从护士鲨 DNA 中分离出的 PCR 产物与哺乳动物 RAG-1 基因具有序列相似性，将用于筛选基因组文库。 由于哺乳动物、鸟类和两栖动物的 RAG-1 和 RAG-2 紧密相连，因此鲨鱼的同源物也可能如此。 将使用 PCR、异源探针或通过搜索开放阅读框来分析含有鲨鱼 RAG-1 的基因组克隆的连接 RAG-2 同源物。 在拟议的规划期之后，鲨鱼 RAG-1 和 RAG-2 同源物将用于通过 Northern 印迹和原位杂交分析不同年龄护士鲨淋巴器官中的表达。 将使用人工底物在哺乳动物成纤维细胞中评估鲨鱼 RAG 蛋白的重组活性。 高等脊椎动物的免疫系统是一个由细胞、器官和组织组成的复杂网络。 这种紧密相连的系统的复杂运作和演变尚不清楚。 免疫功能所需的基本过程是亚基因元件的重排以形成功能性抗原受体。 阐明与哺乳动物有远缘关系的生物体的机制对于我们理解适应性免疫的进化至关重要。 对于拟议的研究计划，选择鲨鱼是因为它们是与已记录适应性免疫系统的哺乳动物关系最远的分类单元。