Factors Involed in Chloroplast RNA Editing

叶绿体 RNA 编辑涉及的因素

• 批准号: 6700712
• 负责人: CARLA E HEGEMAN
• 金额: $ 4.64万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-15 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6700712
• 关键词: DNA directed RNA polymerase; RNA binding protein; SDS polyacrylamide gel electrophoresis; antisense nucleic acid; autoradiography; cell transformation; chloroplasts; computer data analysis; cytidine; denaturing gradient gel electrophoresis; gel mobility shift assay; gene expression; genetically modified plants; intermolecular interaction; ion exchange chromatography; nucleic acid hybridization; nucleic acid structure; plant genetics; polymerase chain reaction; posttranscriptional RNA processing; regulatory gene; tobacco; uridine; yeast two hybrid system

DESCRIPTION (provided by applicant) The long-term objective of this project is to define RNA:RNA and RNA:protein interactions involved in RNA editing. In humans, editing is critical for RNA processing in the expression of several disease-related genes including the Wilm?s tumor susceptibility gene, glutamate receptor subunit B, and apolipoprotein B. This research will elucidate the factors that direct cytidine to uridine editing and increase our understanding of organellar editing mechanisms. During the fellowship period, transgenic and biochemical techniques will be used to identify molecules that interact with the chloroplast rpoB-I editing site. To study the role of RNA secondary structure in editing, rpoB-I will be fused to antisense sequences and expressed in transgenic tobacco chloroplasts. Electrophoretic mobility-shift assays will be performed to identify chloroplast proteins with binding specificity for in vitro transcribed rpoB-I editing sites. The nature of the RNA:protein interaction will be examined using modified editing sequences. The purification and characterization of putative editing proteins will be facilitated by an affinity approach. A functional screen will be utilized for the isolation of genes encoding editing factors.

描述（由申请人提供） 该项目的长期目标是定义RNA：RNA和RNA：蛋白质 RNA编辑涉及的相互作用。在人类中，编辑对于RNA至关重要 在表达几种与疾病相关基因的表达中的处理 Wilm的肿瘤敏感性基因，谷氨酸受体亚基B和 载脂蛋白B。这项研究将阐明指导胞苷的因素 尿疗编辑并增加我们对细胞器编辑的理解 机制。 在研究金期间，转基因和生化技术将是 用于鉴定与叶绿体RPOB-I编辑相互作用的分子 地点。为了研究RNA二级结构在编辑中的作用，RPOB-I将是 融合到反义序列中，并在转基因烟草叶绿体中表达。 将进行电泳移动迁移测定，以鉴定叶绿体 具有结合特异性的蛋白质，用于体外转录的RPOB-I编辑 站点。 RNA的性质：蛋白质相互作用将使用 修改的编辑序列。推定的纯化和表征 编辑蛋白将通过亲和力方法来促进。功能 屏幕将用于隔离编辑因子的基因。