Tools for gene editing in marmosets

绒猴基因编辑工具 总结 本提案的目标是创建一套有效的新工具和技术， 适合用作脑生理学和功能模型的遗传修饰的绒猴。发展 在物种中进行基因操作的有效方法将需要基因组编辑技术的改进， 育种技术、促排卵和取卵、胚胎培养和植入前筛查 进行所需的基因改造。一个大的绒猴群体将是必要的，沿着， 完善技术的基础设施和具有绒猴护理专业知识的跨学科团队， 生殖技术，Cre-drivers和CRISPR技术的组织特异性表达， 测试和神经精神分析本建议的具体目标是：（1）改进技术， 研究绒猴的体外遗传操作，探索绒猴基因编辑的新方法;（2）鉴定安全的 携带基因座并使用用于普遍表达的启动子建立Cas9、Cre和GCaMP绒猴系， 在脑中选择性表达，或区域或细胞类型特异性表达;和（3）产生绒猴模型 与人类大脑疾病的神经生物学相关的基因改变。这些研究将提供工具 对于未来在绒猴中进行的基因编辑实验，将建立一套初始的转基因工具系， 并将为遗传改变的绒猴模型提供初步的原理验证线 可能与人类大脑疾病的神经生物学有关。

Tools for Gene Editing in Marmosets SUMMARY The goal of this proposal is to create an efficient new set of tools and techniques for the generation of genetically modified marmosets suitable for use as models of brain physiology and function. Development of efficient methods of genetic manipulation in the species will require refinement of genome editing technologies, breeding technologies, ovarian stimulation and oocyte retrieval, embryo culture, and pre-implantation screening of embryos for the desired genetic alternations. A large colony of marmosets will be necessary, along with an infrastructure to refine technologies and an interdisciplinary team with expertise in marmoset care, assisted reproduction technologies, tissue-specific expression with Cre-drivers and CRISPR technologies, and behavior testing and neuropsychiatric analysis. The Specific Aims of this proposal are to: (1) Improve technology for in vitro genetic manipulation of marmosets and explore novel methods for marmoset gene editing; (2) Identify safe harbor loci and establish Cas9, Cre, and GCaMP marmoset lines using promoters for ubiquitous expression, selective expression in brain, or regional or cell-type specific expression; and (3) Generate marmoset models with genetic alterations relevant to the neurobiology of human brain diseases. These studies will provide tools for future gene editing experiments in marmosets, will establish an initial set of genetically modified tool lines for use in the field, and will provide initial proof-of-principle lines for marmoset models of genetic alterations potentially relevant to the neurobiology of human brain diseases.