Region Based Functional Annotation of the Mouse Genome

小鼠基因组基于区域的功能注释

• 批准号: 6544478
• 负责人: Timothy Paul O'Brien
• 金额: $ 33.11万
• 依托单位: JACKSON LABORATORY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6544478
• 关键词: artificial chromosomes; biotechnology; computer assisted sequence analysis; cytogenetics; developmental genetics; embryonic stem cell; functional /structural genomics; gene expression; gene mutation; genetic mapping; green fluorescent proteins; laboratory mouse

DESCRIPTION (provided by applicant): The goals of this research project are to identify genes required for mammalian development and to investigate the functional content of the chromosomal region defined by the piebald deletion complex. As genomic sequence information is compiled, the structural and regulatory organization of the genes identified will need to be linked with biological function. Mutagenesis screens, relying on phenotype-driven detection of genetic variation, provide an opportunity to establish this connection. In this proposal we will combine genetic studies with computational and comparative genomic sequence analysis to functionally annotate a defined genomic region. The piebald deletion complex is a collection of overlapping deficiencies that encompass a 15-18.5 cM/-39 Mb interval of distal mouse chromosome 14. Several loci important for development have been mapped within this region, resulting in the characterization of mouse models for understanding gastrulation, skeletal patterning and crarnofacial defects, spinal cord malformation and respiratory distress at birth. Computational and comparative genomic analysis to identify genes and additional conserved sequence features will be merged with functional studies. A traditional two-generation regionally directed ENU mutagenesis screen that takes advantage of the piebald deletions and coat color spotting will be used to recover recessive mutations that are essential for development. In addition, we will develop and employ a novel EGFP-based reporter system that takes advantage of the enhanced green fluorescence protein (EGFP) and color variants to conduct a one generation genetic screen that incorporates the recently developed protocols for embryonic stem cell mutagenesis. This approach offers a higher mutation frequency, the use of a wider variety mutagens, and instant visual classification of all screen-derived progeny (mice and embryos) facilitating in depth phenotype analysis. The mutations recovered in the screen will be fine mapped using the deletion panel in order to identify the gene associated with the mutant phenotype. Together, these approaches will enrich the mouse mutant resource and provide new insights relating biological function with chromosome content and organization.

描述（由申请人提供）：本研究项目的目标是鉴定哺乳动物发育所需的基因，并研究由花斑缺失复合体定义的染色体区域的功能内容。随着基因组序列信息的汇编，所识别基因的结构和调控组织将需要与生物功能联系起来。突变筛选依赖于遗传变异的表型驱动检测，为建立这种联系提供了机会。在这个建议中，我们将结合联合收割机遗传学研究与计算和比较基因组序列分析功能注释一个定义的基因组区域。 花斑缺失复合体是重叠缺陷的集合，其涵盖小鼠14号染色体远端的15-18.5 cM/-39 Mb间隔。几个重要的基因座的发展已被映射在这个区域内，从而在小鼠模型的特点，了解原肠胚形成，骨骼图案和crarnofacial缺陷，脊髓畸形和呼吸窘迫出生。计算和比较基因组分析，以确定基因和其他保守的序列特征将与功能研究合并。传统的两代区域定向ENU诱变筛选利用花斑缺失和毛色斑点，将用于恢复对发育至关重要的隐性突变。此外，我们将开发和采用一种新的基于EGFP的报告系统，该系统利用增强型绿色荧光蛋白（EGFP）和颜色变体进行一代遗传筛选，该筛选结合了最近开发的胚胎干细胞诱变方案。这种方法提供了更高的突变频率，更广泛的诱变剂的使用，以及所有筛选衍生后代（小鼠和胚胎）的即时视觉分类，有助于深入的表型分析。将使用缺失组对筛选中回收的突变进行精细作图，以鉴定与突变表型相关的基因。总之，这些方法将丰富小鼠突变体资源，并提供有关染色体内容和组织的生物功能的新见解。