MAPPING CENTROMERIC REGIONS OF HUMAN CHROMOSOMES

绘制人类染色体着丝粒区域图

基本信息

批准号：
2208499
负责人：
Huntington F Willard
金额：
$ 41.35万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1989
资助国家：
美国
起止时间：
1989-07-01 至 1996-08-31
项目状态：
已结题

项目摘要

The centromeric regions of human chromosomes are dominated by a class of tandemly repeated DNA, alpha satellite, which consists of an extensive group of related, highly diverged repeats, based on a monomer repeat length of ~171 basepairs. Long tandem arrays of alpha satellite, estimated to be hundreds to thousands of kilobasepairs long, are located at the centromeric region of each chromosome. In some cases, centromeric regions are characterized by multiple, independent alpha satellite subsets or by subsets of different classes of satellite DNA (beta or classical), in addition to alpha satellite. Because conventional strategies for mapping human chromosomes cannot easily accommodate long, megabase stretches of repetitive DNA, a directed approach is required for high-resolution physical and genetic mapping of the centromeric region of each human chromosome, for full integration into the complete chromosome maps being generated in other laboratories. The current proposal is an extension of our centromere mapping project, which has been ongoing for the past two years. We propose: (i) to isolate representative DNA probes for the alpha, beta, and classical satellite classes located at or near the centromere of all 24 types of human chromosome and to develop specific sequence tagged sites (STS's) for these centromeric loci; (ii) to develop high-frequency polymorphisms for each centromeric locus and to genotype the 40 CEPH families to contribute to centromere-based genetic linkage maps of each chromosome; (iii) to measure the size and amount of variation of centromeric DNA for each chromosome in a representative collection of individuals to provide a physical estimate to close the "centromere gap" in conventional maps of each chromosome; and (iv) to characterize in detail the complete arrays of four centromeres selected as models (chromosomes 4,7,17, and X), using one- and two-dimensional long-range pulsed-field mapping and short- and long-range cloning in lambda phage and in yeast artificial chromosomes, and strategies designed to identify and clone the junctions between the edges of the centromere array and the chromosome arms. The proposed studies address directly several of the Five-Year Goals of the Human Genome Project: to complete a fully connected human genetic map by providing a polymorphic marker, identified by an STS, at the centromere of each chromosome; to contribute to a fully assembled STS physical map of each human chromosome by providing a well-characterized STS at the centromere of each chromosome and by generating the pulsed-field restriction mapping data needed to fully integrate the centromere into complete chromosome maps; and to generate overlapping sets of closely spaced, ordered markers that span the centromeres of selected chromosomes, with continuity over several million basepairs.

人类染色体的中心层面由一类串联重复的DNA，α卫星，由广泛的基于单体重复长度，相关的高度分化重复序列〜171底面。长长的α卫星阵列，估计为数百至数千千层面长，位于丝粒每个染色体的区域。在某些情况下，中心区域是以多个独立的α卫星亚集或以不同类别的卫星DNA（Beta或经典）的子集，附加α卫星。因为映射的传统策略人类染色体不能轻易容纳长时间的巨型兆瓦重复的DNA，高分辨率需要一种定向方法每个人的中心层面区域的物理和遗传图染色体，以完全集成到完整的染色体图中在其他实验室产生。当前的提议是我们的Centromere映射项目，过去两个年。我们建议：（i）分离代表性的DNA探针位于或附近的Alpha，Beta和古典卫星课程所有24种类型的人类染色体的丝粒，并开发特定这些centromeric基因座的序列标记位点（STS）；（ii）发展每个丝粒基因座的高频多态性和基因型 40个CEPH家族为基于Centromere的遗传连锁图做出贡献每个染色体；（iii）测量变异的大小和量代表性集合中每个染色体的丝粒DNA 个人提供物理估计，以缩小在每个染色体的常规地图；（iv）详细描述选择作为型号（染色体）的四个中心粒的完整阵列 4,7,17和x），使用一维远距离脉冲场在lambda噬菌体和酵母中的映射和短期克隆人造染色体以及旨在识别和克隆的策略 Centromere阵列和染色体边缘之间的连接武器。拟议的研究直接解决了五年目标的几个人类基因组项目：完成完全连接的人类遗传通过提供由STS识别的多态性标记的地图每个染色体的中心；为完全组装的ST做出贡献通过提供良好的STS，每个人类染色体的物理图在每个染色体的中心仪上，并通过产生脉冲场将centromere完全集成到中所需的限制映射数据完整的染色体图；并生成紧密的重叠集间隔，有序的标记，跨越了选定染色体的中心粒，连续性超过数百万底座。