Development of a Diversity Outcross ES Cell Panel for In Vitro Genetics of Toxico

描述(由申请人提供)：菌株背景会强烈影响动物毒性试验的结果。时间和成本的限制排除了体内方法来解决动物模型中缺乏遗传多样性的长期担忧。体外平台在了解遗传背景在毒理学测试中的作用方面要高效得多，而且不会牺牲动物。在这个项目中，我们建议建立一个遗传多样性的大型小组 来自小鼠多样性异交系的ES细胞。DO小鼠是协作杂交所用的8个方正品系的高级杂交后代。DO菌株几乎捕获了实验室菌株中的所有序列变体，并且比F2小鼠多出数百到数千个重组断裂点。它们非常适合于促进复杂的性状定位。ES细胞提供了极大的灵活性，也是体外遗传学平台的理想选择。 ES细胞系使基因组永生，创造了一种可再生资源。原则上，ES细胞可以通过定向分化提供几乎任何类型的细胞。在这种情况下，DOES小组作为一种永久性的、可再生的资源是非常多才多艺的。在第一阶段，我们将通过获得90个独立的、经过验证的DO ES系来确定可行性，并估计在第一阶段小组中观察到的15种模型有毒化合物的细胞毒性的遗传成分。在第二阶段，我们将完成600个ES线面板，使用1400个参考化合物集来鉴定对细胞毒性反应具有显著遗传成分的化合物，并在所鉴定的化合物中定位与该遗传成分有关的基因座。 公共卫生相关性：长期以来，人们对遗传学如何影响毒理学感兴趣，但没有成本效益高、高度可扩展的工具来研究这种关系。该项目将通过开发一个高通量的体外平台来研究毒物暴露的不同反应的遗传基础，从而改善人类健康，降低环境毒物的风险。

DESCRIPTION (provided by applicant): Strain background can strongly influence the outcome of toxicity tests in animals. Time and cost constraints preclude in vivo approaches to longstanding concerns about the lack of genetic diversity in animal models. An in vitro platform is vastly more efficient for understanding the role of genetic background in toxicology testing, and sacrifices no animals. In this project we propose to build a large panel of genetically diverse ES cells from the Diversity Outcross line of mice. The DO mice are an advanced intercross of the 8 founder strains used for the Collaborative Cross. The DO strains capture almost all the sequence variants in laboratory strains, and harbor hundreds to thousands more recombinational breakpoints than F2 mice. They ideally suited for facilitate complex trait mapping. ES cells offer tremendous flexibility and is also ideal for an in vitro genetics platform. DO ES cell lines immortalize DO genomes, creating a renewable resource. In principle, ES cells can provide access to almost any cell type by directed differentiation. In this way, the DO ES panel is extremely versatile as a permanent, renewable resource. In Phase I, we will establish feasibility by deriving 90 independent, validated DO ES lines and estimate the genetic component of cytotoxicity observed for 15 model toxicant compounds in the Phase I panel. In Phase II we will complete the 600 ES line panel, identify compounds with a significant genetic component to cytotoxic response using a 1400 reference compound set, and map the loci mediating that genetic component in the identified compounds. PUBLIC HEALTH RELEVANCE: There is a longstanding interest in how genetics impacts toxicology, but cost effective, highly scalable tools to investigate this relationship were not available. This project will improve human health and reduce the risk of environmental toxicants by developing a high throughput in vitro platform for investigating the genetic basis of variable response to toxicant exposure.