DNA LINKAGE STUDIES OF DEGENERATIVE RETINAL DISEASES

这项建议是为继续我们的DNA连锁研究提供资金 患有常染色体等退行性视网膜疾病的家庭 显性视网膜色素变性(ADRP)和Usher综合征。这个 目标是为这些基因找到紧密连锁的DNA标记 疾病，将疾病基因座映射到特定的染色体区域 并使用这些连锁的DNA标记来描述疾病的特征 精神错乱。 1986年，国家视网膜色素变性基金会成立 一个促进DNA连锁研究的合作项目 视网膜色素变性(RP)。戴格博士是 这个节目。该计划的目标是识别和 确定合适的RP家系特征，以送来家系血样 用于人类遗传突变细胞库的制备和 转化的细胞系的存储，并提供这些细胞系 送到戴格博士的实验室进行准备、测试和处理 DNA的基因。目前，RP收集包含91个细胞系，来自 一个大型ADRP家族和77条来自扩展Usher‘s的线路 综合症家族。集合应该增加50到100行 1988年。我们已经从其中的大多数准备并测试了DNA 样本，并已从其他RP获得额外91个DNA 家人。 这项建议是针对该计划的研究部分，即 DNA连锁检测和数据分析。除 我们和其他人在这些方面测试了经典的遗传标记 家人们，我们已经测试了超过22个DNA探针 家人。我们建议测试150到200个人，测试20个新的 每年使用常规的Southern凝胶分析。我们会 还开发和应用了一种新的、高效的程序来 基于DNA的人类DNA“微型-VNTR”多态检测 采用聚合酶链式反应(PCR)方法进行扩增。 目前有数百种多态DNA探针可供选择 来自科学界，我们假设类似的数字 的迷你汽车将会被发现。 方法有组织培养、基因组DNA提取、制备 DNA探针的Southern凝胶分析、DNA多态性检测 通过聚合酶链式反应扩增，两点和多点连锁 分析。当链接建立后，我们将测试其他 并将使用分子方法来改进信息 标记的内容，开发更紧密的标记并建立 侧翼记分器。ADRP与Usher综合征的连锁标志物 将对早期诊断、基因检测有价值 异质性，最终，在隔离和表征 突变基因。

This proposal is for funds to continue our DNA linkage studies of families with degenerative retinal diseases such as autosomal dominant retinitis pigmentosa (ADRP) and Usher's syndrome. The objectives are to find tightly linked DNA markers for these diseases, to map the disease loci to specific chromosomal regions and to use these linked DNA markers in characterizing the disease loci. In 1986 the National Retinitis Pigmentosa Foundation established a collaborative program to facilitate DNA linkage studies of retinitis pigmentosa (RP). Dr. Daiger is Program Coordinator for this program. The aims of the program are to identify and characterize appropriate RP families, to send family blood samples to the Human Genetic Mutant Cell Repository for preparation and storage of transformed cell lines, and to provide these cell lines to Dr. Daiger's laboratory for preparation, testing and handling of DNAs. At present the RP collection contains 91 cell lines from one large ADRP family and 77 lines from an extended Usher's syndrome family. The collection should increase by 50 to 100 lines in 1988. We have prepared and tested DNAs from most of these samples and have acquired 91 additional DNAs from other RP families. This proposal is for the research component of the program, that is, DNA linkage testing and data analysis. In addition to the classical genetic markers we and others have tested in these families, we already have tested over 22 DNA probes in selected families. We propose to test 150 to 200 individuals with 20 new probes per year using conventional Southern gel analysis. We will also develop and apply a novel, high efficiency procedure for detecting "mini-VNTR" polymorphisms in human DNA, based on DNA amplification using the polymerase chain reaction (pcr) method. There are hundreds of polymorphic DNA probes currently available from the scientific community and we hypothesize that like numbers of mini VNTRs will be found. Methods are tissue culture, extraction of genomic DNAs, preparation of probe DNAs Southern gel analysis, detection of DNA polymorphisms by pcr amplification, and two point and multipoint linkage analysis. When linkage is established we will test additional families and will use molecular methods to improve the information content of the marker, develop closer markers and establish flanking markers. Linked markers for ADRP and Usher's syndrome will be of value in early diagnosis, in detection of genetic heterogeneity and eventually, in isolation and characterization of the mutant genes.