EXPRESSION AND LOCATION OF DNA SEQUENCES ON CHROMOSOME 21

我们建议使用重复序列来寻找 参与与其他疾病的DS相关的病理 还探讨了受21号染色体影响的调控机制 重复序列在这些基因表达中的作用 大脑中的其他基因。计划进行以下实验： 1.利用克隆的重复序列作为遗传标记 鉴定和分离21号染色体上的异常DNA片段 定义缺失、移位和其他重排 在唐氏综合征和其他患者中观察。 2.特定目的1中分离的DNA片段的特征。 测序分析将揭示任何开放的阅读框架以及 基因被扩增。进行染色体作图以定位 21号染色体上的这样的序列。 3.基因序列在RNA和蛋白质水平的表达。我们 应使用在特定目标#1中获得的基因序列以及 已有的胎儿脑cDNA文库中的克隆映射到 21号染色体。组织中RNA的原位杂交 16三体小鼠胎脑切片的比较 将识别基因序列，揭示 优先杂交到一个或另一个。 这些序列和其他序列将被亚克隆到表达载体中 来自这样的重组子的蛋白质产物将被注射 进入老鼠体内以产生抗体。这样的免疫荧光 与三体胎儿脑组织切片结合的抗体 16只小鼠或来自正常小鼠的小鼠将定量显示或 定性地，这些基因序列是否被表达 在胚胎16三体小鼠的大脑中有不同的表现。 4.研究人类特有的重复序列 出现在21号染色体上。特定于该基因的重复序列 人类物种将从21号染色体基因组文库中分离出来 通过差异杂交。他们的监管角色将是 特别是在它们对基因表达的控制方面进行了探索 大脑。

We propose to use repetitive sequences in search of genes that are involved in the pathology associated with DS on other diseases affected by chromosome 21 and also to explore the regulatory function of repetitive sequences in the expression of these and other genes in the brain. The following experiments are planned: 1. Use of cloned repetitive sequences as genetic markers to identify and isolate abnormal DNA fragments along chromosomal 21 defining deletions, translocations and other rearrangements observed in Down syndrome and other patients. 2. Characterization of DNA fragments isolated in specific aim #1. Sequencing analysis will reveal any open reading frames and whether gene is amplified. Chromosomal mapping is carried out to locate such sequences on chromosome 21. 3. Expression of gene sequences at the RNA and protein level. We shall use gene sequences obtained in specific aim #1 as well as existing cDNA clones from fetal brain cDNA library mapped to chromosome 21. In situ hybridization to RNA in tissue section of fetal brain from trisomy 16 mouse as compared to that of normal mouse will identify gene sequences that reveal preferential hybridization to one or the other. Such sequences and others will be subcloned into expression vector and the protein product from such recombinants will be injected into mice for antibody production. Immunofluorescence of such antibody which binds to tissue section of fetal brain of trisomy 16 mouse or that from normal mouse will reveal quantitatively or qualitatively whether these gene sequences are expressed differently in the brain of a fetal trisomy 16 mouse. 4. Study of repetitive sequences that are human specific and are present on chromosome 21. Repetitive sequences specific to the human species will be isolated from chromosome 21 genomic library by differential hybridization. Their regulatory role will be explored especially in their control of expression of genes In the brain.