IOS EDGE: Expanding the toolkit for functional genetics in threespine stickleback to place genomics into its natural context

为了了解人类生物学，研究人员使用非人类物种作为替身或“模型”。因为没有一个单一的物种是人类生物学的完美模型，科学受益于广泛的模式生物组合，每一个都与人类生物学有自己的相关性。这个项目将创造新的方法来研究一个新兴的模型，三棘棘鱼（Gasterosteus aculeatus）。与传统的实验室模型相比，这种小鱼具有重要的优势。自从大约12000年前北半球冰川消退以来，许多有益的基因变异在数千个不同的自然种群中幸存下来。与实验室遗传筛选不同，这些自然变异揭示了生物创新：例如，抵抗寄生虫的新方法。实验室遗传筛选揭示了导致异常发育或生理的基因突变。一个庞大的研究团体（100个实验室）正在研究棘鱼，以了解其行为、发育和免疫学。研究小组已经确定了影响大脑和骨骼发育等重要特征的候选基因，但需要通过实验改变基因功能的工具来证实。这笔拨款将支持一个由五名研究人员组成的团队为棘鱼开发新的基因编辑方法。因为刺鱼在野外也很容易被研究，这个研究小组可以采取独特的步骤，将实验室遗传学与自然环境中的基因功能联系起来。研究人员将通过新的在线社区和面对面的课程，培训来自世界各地的本科生、研究生和研究人员。该项目还将建设基础设施（一个库存中心），以使研究有用的人口永久化，并为美国土著学生和社区提供研究经验，使他们接触到对当地土著动物的科学研究。棘鱼在生物生物学中是一个领先的模型，原因如下：海洋种群独立地殖民了许多不同的沿海流域，从大规模复制的自然实验中产生了异常的表型变异。2. 祖先的等位基因在现代海洋种群中持续存在，使人们能够在已知的生态环境中绘制适应性性状的遗传图谱。3. 在实验室和自然界中，刺鱼很容易大量繁殖和操纵。4. 已发表的基因组、转录组和微生物组为强有力的候选基因鉴定和探针开发提供了参考。因此，数百名研究人员研究了刺鱼适应的遗传学。然而，这方面的研究主要局限于相关的方法：QTL定位、散度定位和关联研究。研究人员很少使用实验性基因操作来验证可疑的候选基因。为了实现棘鱼的功能基因组研究，该项目将改进基因编辑方法。首先，研究小组将使用转录组学和ChIP-Seq来改进基因组注释，以更好地定位候选基因。其次，他们将开发更有效的基因编辑方法，通过胚胎显微注射或细胞培养转染，或使用病毒介导的基因转移来传递CRISPR-Cas9。最后，该团队将建立一个库存中心，在那里可以维护自然和转基因系，并将其运送给用户，从而能够对跨越棘鱼的标准化遗传背景进行研究。自然的多样性。这项工作将通过网络研讨会、方法博客、学生交叉培训和大型实验课程进行传播。这将为世界各地的本科生、研究生、博士后和同事以及阿拉斯加的美洲原住民社区提供研究培训。

'To learn about human biology, researchers use non-human species as stand-ins, or 'models'. Because no single species is a perfect model of human biology, science benefits from a broad portfolio of model organisms, each with its own relevance to human biology. This project will create new methods to study an emerging model, the threespine stickleback (Gasterosteus aculeatus). This small fish offers an important advantage over traditional lab models. Since the glaciers retreated in the northern hemisphere ~12,000 years ago, many beneficial genetic variants have survived in thousands of distinct natural populations. Unlike laboratory genetic screens, which reveal mutations in genes causing abnormal development or physiology, these natural variants reveal biological innovations: new ways of resisting parasites, for instance. A large community of researchers (100 labs) now studies stickleback to understand behavior, development, and immunology. The research team has identified candidate genes affecting important traits like brain and bone development, but require tools to experimentally alter gene function for confirmation. This grant will support a team of five researchers developing new gene-editing methods for stickleback. Because stickleback are easily studied in the wild as well, this research team can take the unique step of linking laboratory genetics back to gene function in their natural environment. The researchers will train undergraduates, graduate students, and researchers from around the world, via new online communities and in-person courses. The project will also build infrastructure (a stock center) to perpetuate research-useful populations and provide research experiences for Native American students and communities, exposing them to scientific studies of local native animals.Stickleback are a leading model in organismal biology for several reasons: 1. Marine populations independently colonized many diverse coastal watersheds, yielding exceptional phenotypic variation from a massively replicated natural experiment. 2. Ancestral alleles persist in modern oceanic populations, enabling genetic mapping of adaptive traits in known ecological contexts. 3. Stickleback are easy to breed and manipulate in large numbers in the lab and in nature. 4. Published genomes, transcriptomes, and microbiomes provide references for robust candidate gene identification and probe development. Consequently, hundreds of researchers study the genetics of stickleback adaptation. However, this research is largely confined to correlative approaches: QTL mapping, divergence mapping, and association studies. It remains rare for researchers to validate suspected candidate genes using experimental gene manipulation. To enable functional genomic studies of stickleback, this project will refine methods for gene editing. First, the research team will use transcriptomics and ChIP-Seq to improve genome annotation, to better locate candidate genes. Second, they will develop methods for more efficient gene editing with CRISPR-Cas9 delivered via embryo microinjection or cell culture transfection, or using viral-mediated gene transfer. Finally, the team will construct a stock center where natural and transgenic lines can be maintained and shipped to users, enabling studies on standardized genetic backgrounds spanning sticklebacks? natural diversity. The work will be disseminated via webinars, methods blogs, cross training of students, and a large lab course. This will provide research training to undergraduates, graduate students, postdocs, and colleagues around the world, as well as Native American communities in Alaska.