SATURATED GENOME ANALYSIS VIA 2D GELS

通过 2D 凝胶进行饱和基因组分析

基本信息

批准号：
2209084
负责人：
CHARLES L SIDMAN
金额：
$ 15.59万
依托单位：
UNIVERSITY OF CINCINNATI
依托单位国家：
美国
项目类别：
财政年份：
1992
资助国家：
美国
起止时间：
1992-06-08 至 1995-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2209084
关键词：
computer assisted sequence analysis gel electrophoresis genetic mapping genetic markers genetic techniques genome nucleic acid repetitive sequence polymerase chain reaction sequence tagged sites southern blotting

项目摘要

This application is directed at "Pilot Projects or Feasibility Studies for Genomic Analysis" (PA-90-21) in support of the Human Genome Initiative (HGI), and seeks to develop two-dimensional (2D) gels of DNA fragments as a method to detect and resolve up to several thousand independent genomic segments in a single gel. Such an ability would enable at least an order- of-magnitude improvement in the rapidity of producing saturated genetic and physical maps of any organism, and would greatly facilitate multiple types of genomic analysis through the ability to simultaneously examine the segregation or other status of large numbers of markers distributed across an entire genome. The proposed study will employ the mouse as its experimental mammalian model due to the wealth of available inbred and well-characterized genetic stocks; however, the methods developed should be generally applicable with little further modification to any genetic system. In brief, existing techniques of 2D electrophoresis of DNA fragments (by size and base composition/sequence), probing for highly repetitive DNA sequences, computer-assisted analysis of complex spot patterns and classical mouse genetics will be combined in a novel synthesis. The repetitive sequences to be explored will include both previously characterized repeats such as Intracisternal A Particles (IAPs), which are widely distributed throughout the mouse genome and are highly polymorphic both within and between inbred strains, and oligonucleotides representing subsets of LINE-1 elements. A single blot probed with one of these sequences should enable simultaneous analysis of hundreds or even thousands of independent genomic segments. Such a blot could be reprobed as desired to reveal the distribution of additional families of repetitive sequences, and individual spots could be recovered for analysis of unique flanking regions and determination of a generally accessible Sequence Tagged Site (STS) from each resolved copy of a repetitive sequence.

该应用针对“试点项目或可行性研究基因组分析“（PA-90-21）支持人类基因组倡议（HGI），并试图开发DNA片段的二维（2D）凝胶作为一个检测和解决多达数千种独立基因组的方法单个凝胶中的片段。这样的能力至少可以订购 - 产生饱和遗传和任何有机体的物理地图，并且将极大地促进多种类型通过同时检查的能力来进行基因组分析分布的大量标记的隔离或其他状态整个基因组。拟议的研究将使用鼠标作为其由于可用的近交和实验性哺乳动物模型特征良好的遗传库存；但是，开发的方法应该是通常适用于任何遗传的进一步修改系统。简而言之，DNA的2D电泳现有技术片段（按大小和基础组成/序列），探测高度重复的DNA序列，复杂点的计算机辅助分析图案和经典的小鼠遗传学将结合在小说中合成。要探索的重复序列将包括先前表征的重复，例如肠内A颗粒（IAP），广泛分布在整个小鼠基因组中，高度多态性在近交菌株和寡核苷酸之间的多态性代表行1元素的子集。一个印迹，用一个这些序列应同时分析数百甚至成千上万的独立基因组段。这样的印迹可以被重新发行根据需要揭示重复的其他家族的分布序列和单个斑点可以回收以分析独特侧翼区域和确定通常可访问的序列来自重复序列的每个已解决副本的标记位点（STS）。