Streamlined cloning of auditory and vestibular mutants by whole genome sequencing

通过全基因组测序简化听觉和前庭突变体的克隆

基本信息

批准号：
8411127
负责人：
SEAN G MEGASON
金额：
$ 20.13万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-01-10 至 2014-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8411127
关键词：
Adult Affect Alleles Animal Model Applied Genetics Area Auditory Base Sequence Bioinformatics Biological Chemicals Cloning Computer software DNA Resequencing Defect Development Ear Embryo Embryonic Development Equilibrium Future Gamma Rays Genes Genetic Genetic Polymorphism Genetic Screening Genome Genomics Goals Hair Health Hearing Histocompatibility Testing Human Human Development Individual Insertional Mutagenesis Investments Labyrinth Learning Maps Methods Modeling Molecular Molecular Analysis Morphogenesis Mutagenesis Mutagens Mutate Mutation Natural regeneration Organ Pathway interactions Phenotype Positioning Attribute Publishing Reaction Research Research Personnel Semicircular canal structure Structure Technology Time Vertebrates Zebrafish base cell type cellular transduction cost deafness feeding gene function genetic linkage analysis genome sequencing model organisms databases mutant next generation sequencing novel positional cloning prevent research study sound

项目摘要

DESCRIPTION (provided by applicant): Over the last two decades zebrafish has emerged as one of the preeminent model organisms. This rise was in large part due to the promise of using forward genetic screens in a vertebrate animal to discover and elucidate the function of genes relevant to human development and health. Numerous genetic screens have now been performed both for general embryonic morpohological phenotypes as well as more specialized functional screens such as for balance and hearing. These screens have been very successful in identifying mutants and there are now over 6000 described chemically induced mutant lines in zebrafish. However, the gene mutated in these lines has only been identified in less than half the cases due to the current difficulty of positional cloning. The high cost and labor currently required for positional cloning prevents a molecular analysis of many current mutants as well as discouraging future genetic screens. Recent advances in next generation sequencing have reduced the cost of sequencing by several orders of magnitude such that it is now possible to routinely resequence entire genomes. These advances have already revolutionized many areas of genomics yet the way people do positional cloning in zebrafish and other model organisms has changed relatively little. Here we propose to apply next generation sequencing technologies to streamline positional cloning of mutants in model organisms focusing on zebrafish inner ear mutants. Our goal is to reduce the labor and cost of positional cloning by one to two orders of magnitude. We will compare two different approaches for cloning-by-sequencing based on linkage and homozygosity mapping by cloning 10 existing mutants in two mutant classes-semicircular canal morphogenesis and deafness.

描述（由申请人提供）：在过去的二十年中，斑马鱼已成为杰出的模型生物之一。这种上升在很大程度上是由于有望在脊椎动物中使用正向遗传筛选来发现和阐明与人类发育和健康相关的基因功能。现在已经为一般的胚胎畸形表型以及更专业的功能筛选（例如平衡和听力）进行了许多遗传筛选。这些屏幕在识别突变体方面非常成功，现在有6000多个描述的化学诱导的突变系在斑马鱼中。但是，由于当前的位置克隆难度，这些线中突变的基因仅在不到一半的情况下被鉴定出来。位置克隆目前所需的高成本和劳动力阻止了许多当前突变体的分子分析，并阻止未来的遗传筛查。下一代测序的最新进展将测序的成本降低了几个数量级，因此现在可以常规地重新恢复整个基因组。这些进步已经彻底改变了许多基因组学领域，但是人们在斑马鱼和其他模型生物中进行定位克隆的方式却相对较小。在这里，我们建议将下一代测序技术应用于以斑马鱼内耳突变体的模型生物中突变体的位置克隆。我们的目标是将克隆的劳动力和成本减少一到两个数量级。我们将根据将10个现有突变体在两个突变类圆形管道形态发生和耳聋中克隆出来的连接和纯合性映射来比较两种不同的方法。