Haplotype Sequencing via Single Molecule Hybridization

通过单分子杂交进行单倍型测序

• 批准号: 7140303
• 负责人: Bhubaneswar Mishra
• 金额: $ 28.55万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7140303
• 关键词: alleles; biotechnology; genome; human tissue; molecular biology information system; nanotechnology; nucleic acid hybridization; nucleic acid probes; nucleic acid sequence; restriction mapping; technology /technique development

DESCRIPTION (provided by applicant): We seek funding to develop, within 10 years, a technology to sequence a human size genome of about 6 Gigabases including both haplotypes. We aim to accomplish these goals by successfully integrating three different component technologies: (1) Optical Mapping to create Ordered Restriction Maps with respect to an enzyme, (2) Hybridization of a pool of oiigonucleotide probes (LNA probes) with Single Genomic DNAs on surface, and (3) Algorithms to solve "localized versions" of PSBH (Positional Sequencing by Hybridization) problems over the whole genome. The project is planned in two stages: (1) Pilot Study to Assess Scientific Soundness [R21] and (2) Large-Scale System Engineering [R33]. The R21 phase aims to demonstrate first the soundness of whole-genome mapping of LNA probe hybridization sites, and then algorithmic feasibility of combining these maps into haplotype sequences. The potential for success of these two aims may be inferred from our preliminary work on (1) haplotype mapping of T. pseudonana and a segment of human chromosome 4; (2) fluorescent imaging of DNA and its validation by AFM technology; and (3) existing body of work on optical mapping by our investigators. The R33 phase aims to engineer the final system by constructing in succession: (1) high throughput optical system, (2) preliminary validation by sequencing 100bp segment of P. falciparum genome (small-size), (3) more complex validation by sequencing 100bp segment of H. sapiens genome (large-size), (4) final system engineering and validation by sequencing the entire H. sapiens genome.

描述（由申请人提供）：我们寻求资金在 10 年内开发一项技术，对大约 6 GB 的人类基因组（包括两种单倍型）进行测序。我们的目标是通过成功集成三种不同的组件技术来实现这些目标：(1) 光学作图以创建关于酶的有序限制性图谱，(2) 寡核苷酸探针（LNA 探针）池与表面单基因组 DNA 的杂交，以及 (3) 解决 PSBH（杂交定位测序）问题的“本地化版本”的算法 整个基因组。 该项目计划分两个阶段：(1) 评估科学可靠性的试点研究 [R21] 和 (2) 大规模系统工程 [R33]。 R21阶段的目的是首先证明LNA探针杂交位点全基因组图谱的合理性，然后证明将这些图谱组合成单倍型序列的算法可行性。这两个目标的成功潜力可以从我们的初步工作中推断出来：(1) T. pseudonana 和人类 4 号染色体片段的单倍型作图； (2) DNA荧光成像及其AFM技术验证； (3) 我们的研究人员现有的光学测绘工作。 R33阶段旨在通过连续构建以下方式来设计最终系统：（1）高通量光学系统，（2）通过对恶性疟原虫基因组（小尺寸）的100bp片段进行测序进行初步验证，（3）通过对智人基因组（大尺寸）的100bp片段进行测序来进行更复杂的验证，（4）通过对整个智人进行测序来进行最终系统工程和验证 基因组。