HIGH RESOLUTION GENE MAPPING BY IN SITU HYBRIDIZATION

通过原位杂交进行高分辨率基因图谱

• 批准号: 2838858
• 负责人: DAVID C WARD
• 金额: $ 42.21万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-05-01 至 2000-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2838858
• 关键词: chromosomes; fluorescent in situ hybridization; human genetic material tag; molecular cloning; nucleic acid probes; nucleic acid sequence; plasmids; sequence tagged sites

A physical map of the human genome in terms of a set of overlapping yeast artificial chromosome (YAC) clones is now nearing completion (Chumakov, et al.; Hudson, et al., 1995). One of the necessary prerequisites to initiating a large scale sequencing effort will be to convert the YAC contig(s) into a minimum tiling path of cosmid or PAC clones. The major objective of this project is to streamline the generation of sequencing substrates and to minimize downstream sequencing redundancy by identifying cosmid and PAC clones using multiplex array screening and by using novel optical imaging techniques for constructing contig maps and identifying those clones which exhibit a minimal sequence overlap. The specific goals are: 1) To develop a procedure for screening gridded arrays of cloned DNA with up to 31 probes simultaneously, each identifiable by a unique spectral signature based on fluor- composition, using an epifluorescence microscope based array scanner. 2)To construct a high resolution cosmid and PAC contig map covering the entirety of human chromosome 12. (130Mb). Cosmid and PAC clones, corresponding to a minimal tiling path of nonchimeric YAC clones previously characterized, will be identified by conventional library screening and by multiplex fluorescence imaging. These clones will be ordered and their overlap ascertained by multicolor FISH to YAC DNA that has either been stretched on a glass slide or immobilized in an oriented fashion and extended in a mild electric field. Six or more cosmid or PAC clones, each labeled with a different fluorophores or combinination of fluorophores, will be hybridized simultaneously. 3) To continue to add new markers to our existing YAC contig map and our proposed cosmid(PAC maps of chromosome 12. Our current map consists of 1100 YACs and contains 1000 markers, approximately 1 per 130Kb. These include highly polymorphic genetic markers, monomorphic and anonymous markers as well as genes and ESTs. In order to increase the utility of our map until the complete sequence is obtained, we will use PCR screening and multiplex hybridization to arrayed ESTs to incorporate at least 1000 new markers into the map over the next three years.

人类基因组的物理图谱， 重叠酵母人工染色体（YAC）克隆， 接近完成（Chumakov等人;哈德逊等人，1995年）。一 启动大规模测序的必要前提 将努力将YAC重叠群转换为最小的平铺 粘粒或PAC克隆的路径。该项目的主要目标是 是简化测序底物的产生， 通过鉴定粘粒使下游测序冗余最小化 和PAC克隆使用多重阵列筛选和使用新的 用于构建重叠群图谱的光学成像技术， 鉴定显示最小序列重叠的那些克隆。 具体目标是：1）制定筛选程序 同时具有多达31个探针的克隆DNA的网格阵列， 每一个都可通过基于荧光的独特光谱特征来识别， 组合物，使用基于落射荧光显微镜的阵列 扫描仪2）构建高分辨率的粘粒和PAC重叠群 覆盖了整个人类12号染色体的图谱。（130Mb）。 Cosmid和PAC克隆，对应于 以前表征的非嵌合YAC克隆， 通过常规文库筛选和多重鉴定 荧光成像这些克隆人将被命令和他们的 通过FISH确定与YAC DNA的重叠， 或者在载玻片上拉伸，或者固定在定向的 时尚，并在温和的电场中延伸。六个或更多个粘粒或 PAC克隆，每个用不同的荧光团标记，或 荧光团的组合将同时杂交。 3)为了继续向我们现有的YAC重叠群图谱中添加新的标记， 和我们提出的粘粒（12号染色体的PAC图谱。我们 目前的图谱由1100个YAC组成并包含1000个标记， 大约每130 Kb 1个。其中包括高度多态性 遗传标记、单态和匿名标记以及 基因和EST。为了增加地图的实用性， 获得完整序列后，我们将使用PCR筛选， 与阵列化EST的多重杂交，以掺入至少1000个 在地图上添加新的标记

项目成果

In situ hybridization banding of human chromosomes with Alu-PCR products: a simultaneous karyotype for gene mapping studies.

人类染色体与 Alu-PCR 产品的原位杂交显带：用于基因作图研究的同步核型。

• DOI: 10.1016/0888-7543(91)90374-n
• 发表时间: 1991
• 期刊: Genomics
• 影响因子: 4.4
• 作者: Baldini,A;Ward,DC
• 通讯作者: Ward,DC

CEPH viewer: a client-server database to browse and manipulate CEPH physical mapping and linkage data.

CEPH 查看器：一个客户端-服务器数据库，用于浏览和操作 CEPH 物理映射和链接数据。

• DOI: 10.1016/0888-7543(95)80147-e
• 发表时间: 1995
• 期刊: Genomics
• 影响因子: 4.4
• 作者: Nadkarni,PM;Bray-Ward,P
• 通讯作者: Bray-Ward,P

Evaluation of the utility of interphase cytogenetics to detect residual cells with a malignant genotype in mixed cell populations: a Burkitt lymphoma model.

评估间期细胞遗传学在检测混合细胞群中具有恶性基因型的残留细胞的效用：伯基特淋巴瘤模型。

• DOI: 10.1089/dna.1993.12.637
• 发表时间: 1993
• 期刊: DNA and cell biology
• 影响因子: 3.1
• 作者: Ried,T;Lengauer,C;Lipp,M;Fischer,C;Cremer,T;Ward,DC
• 通讯作者: Ward,DC

Rapid physical mapping of cloned DNA on banded mouse chromosomes by fluorescence in situ hybridization.

• DOI: 10.1016/0888-7543(92)90412-l
• 发表时间: 1992
• 期刊: Genomics
• 影响因子: 4.4
• 作者: Ann L. Boyle;David M. Feltquite;N. Dracopoli;David E. Housman;David C. Ward

Mapping of human microtubule-associated protein 1B in proximity to the spinal muscular atrophy locus at 5q13.

靠近 5q13 脊髓性肌萎缩基因座的人类微管相关蛋白 1B 的定位。

• DOI: 10.1073/pnas.88.17.7873
• 发表时间: 1991
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Lien,LL;Boyce,FM;Kleyn,P;Brzustowicz,LM;Menninger,J;Ward,DC;Gilliam,TC;Kunkel,LM
• 通讯作者: Kunkel,LM