END SEQUENCING AND FINGERPRINTING OF MOUSE BAC LIBRARIES

小鼠 BAC 文库的末端测序和指纹图谱

• 批准号: 6451359
• 负责人: SHAYING ZHAO
• 金额: $ 145.83万
• 依托单位: INSTITUTE FOR GENOMIC RESEARCH
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-30 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6451359
• 关键词: animal data; animal genetic material tag; animal tissue; artificial chromosomes; genetic library; laboratory mouse; nucleic acid sequence; restriction fragment length polymorphism

As the human project shifts into production sequencing, a plan is developing for sequencing the mouse genome. The current plan employs a "sequence first, map second" strategy which, like the human project, is based on shotgun sequencing of BAC clones comprising contiguous regions of the genome of the selected mouse strain (C57BL6/J). Libraries constructed in bacterial artificial chromosomes (BAC) vectors have become the choice for high throughput genomic sequencing projects because of their cloning capacity and high stability. The current whole-genome approach, which has been validated in the human genome sequencing project, uses BAC end sequences and restriction fingerprints to build BAC contigs. Restriction fingerprints not only serve as independent confirmation of BAC contigs assembled based on BAC end sequence, but also enable identification of chimeric BACs and clones with internal deletions. This map-as-you-go strategy saves substantial time and effort in constructing sequence ready maps. This proposal is to perform BAC end sequencing at TIGR and restriction fingerprinting at the Clemson University Genomics Institute Physical Mapping Center from the mouse BAC library or libraries constructed to support mouse genome sequencing. The clones, the sequences, gel images of the fingerprints, and the binning of the clones using the FPC program (C. Soderlund, Sanger Centre) will be an available resource for those sequencing the mouse genome, and the community at large. We will continue our focus on quality in this BAC clone end sequencing and restriction fingerprinting project while developing and implementing automation and new methodologies for reducing costs and increasing throughput.

随着人类项目转向生产测序，一项对小鼠基因组进行测序的计划正在制定中。目前的计划采用了“序列优先，地图第二”的策略，与人类的计划一样，基于对BAC克隆的鸟枪式测序，这些克隆包括所选小鼠品系(C57BL6/J)基因组的连续区域。在细菌人工染色体(BAC)载体中构建的文库因其克隆能力和高稳定性而成为高通量基因组测序项目的选择。目前的全基因组方法已经在人类基因组测序项目中得到验证，它使用BAC末端序列和限制指纹来构建BAC重叠群。限制性内切酶指纹不仅可以作为根据BAC末端序列组装的BAC重叠群的独立确认，而且还可以识别嵌合BAC和带有内部缺失的克隆。这种即用即用映射策略节省了构建序列就绪映射的大量时间和精力。这项提议是在TIGR处进行BAC末端测序，并在克莱姆森大学基因组研究所物理图谱中心从小鼠BAC文库或为支持小鼠基因组测序而构建的文库中进行限制指纹分析。克隆、序列、指纹的凝胶图像以及使用FPC程序(C.Soderlund，Sanger Centre)对克隆进行分类将为那些对小鼠基因组进行测序的人以及整个社区提供可用的资源。我们将在这个BAC克隆末端测序和限制指纹分析项目中继续关注质量，同时开发和实施自动化和新方法，以降低成本和增加产量。