Identification and Functional Analysis of the Mouse Deaf

聋鼠的鉴定及功能分析

• 批准号: 6789016
• 负责人: David C Kohrman
• 金额: $ 3.77万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6789016
• 关键词: cochlea; deafness; electrophysiology; gene expression; gene mutation; genetic mapping; genetically modified animals; human genetic material tag; laboratory mouse; molecular cloning; nucleic acid sequence; protein localization; sensorineural hearing loss

DESCRIPTION: (Adapted from applicant's abstract): The primary objective of this proposal is the identification and characterization of the gene responsible for inherited deafness in the mouse mutant known as 'spinner'. Mutation of this gene results in sensorineural hearing loss similar to that found in many human nonsyndromic deafness disorders. The approaches described will attempt to correlate effects at the single gene level with those occurring at the cellular level and the level of the intact cochlea. A positional cloning strategy will be used to identify the affected gene. Gene localization information will be derived from an existing high resolution genetic cross and a set of genomic DNA clones that span the candidate region. Positional candidate genes mapped to this region in mouse, and to the region of conserved linkage in humans, will be evaluated for mutation in spinner mice. To further narrow the candidate region, genomic DNA clones from the candidate region will be microinjected into sr/sr zygotes. DNA clones that contain a functional version of the normal gene are expected to produce phenotypic correction in the resulting transgenic progeny. This DNA will be directly screened for the presence of the affected gene. The biological role of the spinner gene in the cochlea will be examined using several approaches. High resolution phenotypic analysis of affected mice will be performed to identify early defects in the cochlea that result from mutation of the spinner gene. Sensory cell function in the cochlea will be assessed by measurement of evoked responses in spinner mice. Ultrastructural and immunocytochemical analyses will be performed to detail the degenerative process in the mutant cochlea. Database analysis of the gene's primary sequence will be used to identify related genes and protein motifs that may provide insight into gene function. The expression pattern of the spinner gene, and the subcellular localization of its encoded protein, will be determined. Based upon comparative genetic data, the human version of the spinner gene is a positional candidate for the gene affected in a nonsyndromic deafness disorder, DFNB6. The human gene will be directly evaluated for mutations in individuals with inherited defects at the DFNB6 locus. This project will result in the identification of a critical gene in the mammalian inner ear, provide the basis for a model of this gene's role in the cochlea, and investigate the involvement of the gene in human nonsyndromic hearing loss.

描述：（改编自申请人的摘要）：此的主要目的 提案是负责基因的识别和表征 在称为“旋​​转器”的小鼠突变体中的遗传耳聋。突变 基因导致感觉到听力损失类似于许多人 非呼吸障碍障碍。所述的方法将尝试 在单个基因水平上的效应与在细胞处发生的效应 水平和完整的耳蜗的水平。位置克隆策略将 用于识别受影响的基因。基因定位信息将是 源自现有的高分辨率遗传十字和一组基因组DNA 跨越候选区域的克隆。位置候选基因映射到 小鼠中的该区域以及人类保守联系的区域将是 评估旋转小鼠中突变。为了进一步缩小候选区域， 来自候选区域的基因组DNA克隆将微注射到SR/SR中 Zygotes。包含正常基因功能版本的DNA克隆是 预计将在产生的转基因后代产生表型校正。 该DNA将直接筛选出受影响的基因的存在。这 旋转基因在耳蜗中的生物学作用将使用 几种方法。受影响小鼠的高分辨率表型分析将 进行以确定突变导致的耳蜗中的早期缺陷 旋转基因。耳蜗中的感觉细胞功能将通过 旋转小鼠中诱发反应的测量。超微结构和 将进行免疫细胞化学分析，以详细介绍退行性 突变耳蜗中的过程。基因主要序列的数据库分析 将用于识别可能提供的相关基因和蛋白质基序 洞悉基因功能。旋转基因的表达模式，以及 将确定其编码蛋白的亚细胞定位。基于 比较遗传数据，旋转基因的人类版本是位置 在非综合性耳聋障碍中受影响的基因候选者，DFNB6。这 人类基因将直接评估患有 DFNB6基因座的继承缺陷。这个项目将导致 鉴定哺乳动物内耳中关键基因，提供基础 对于该基因在耳蜗中的作用的模型，并研究了参与 人类非综合性听力损失中的基因。