Mendelian inheritance of artificial chromosomes

人工染色体的孟德尔遗传

• 批准号: 10666591
• 负责人: Ben E. Black
• 金额: $ 116.96万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-10 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10666591
• 关键词: Address; Animal Model; Animals; Area; Artificial Chromosomes; Artificial Mammalian Chromosomes; Behavior; Biological Models; Biotechnology; Cell division; Centromere; Chromatin; Chromosomal Stability; Chromosome Pairing; Chromosomes; Circular DNA; Clinic; Custom; DNA; DNA Sequence; Data; Development; Elements; Embryo; Ensure; Epigenetic Process; Eukaryota; Female; Foundations; Genes; Genetic; Genetic Recombination; Genome; Genome Components; Genome engineering; Germ Cells; Goals; Harvest; Histones; Human Chromosomes; Human Genome Project; In Vitro; Industrialization; Inherited; Investments; Laws; Mammalian Cell; Mammalian Chromosomes; Mammals; Medicine; Meiosis; Mitosis; Mitotic; Mitotic Cell Cycle; Mus; Nucleic Acid Sequence Homology; Nucleosomes; Organ Transplantation; Outcome; Patients; Privatization; Prophase; Research Personnel; Research Project Grants; Saccharomycetales; Sex Chromosomes; Sexual Reproduction; Source; Specific qualifier value; Technology; Testing; Time; Variant; Work; Writing; Yeasts; centromere protein A; design; drug development; frontier; genetic element; genetic payload; in vivo; innovation; insight; mouse model; quantum; segregation; success; synthetic biology; synthetic construct; telomere; tool; transmission process; vector

Synthetic mammalian artificial chromosomes (MACs) represent a new frontier in genome technology, with the potential to transform chromosome and synthetic biology and stimulate the development of numerous radical advances in medicine. Human Genome Project-Write aims to generate an entire set of synthetic human chromosomes. Short of this ambitious goal, MACs have enormous potential for breakthroughs in biotechnology and medicine, such as creating humanized animal models for drug development or for harvesting patient- personalized organs for transplantation. Furthermore, building MACs from minimal components will advance our fundamental understanding of what comprises a mammalian chromosome. As vehicles for genetic inheritance, fully functional chromosomes are faithfully transmitted through mitosis and the specialized meiotic divisions underlying eukaryotic sexual reproduction and Mendelian inheritance. Our goal is to construct the first MACs that achieve faithful inheritance through the germline, using mouse as a model system. One obstacle is the centromere, the locus on each chromosome that directs transmission through both mitosis and meiosis. Because mammalian centromeres are not encoded in the DNA sequence, it is unclear how to build synthetic chromosomes containing this crucial element. There are additional challenges to create MACs that pair and recombine as homologous chromosomes in meiosis. To solve these problems, we will hijack the existing cellular machinery for assembling centromere chromatin and incorporate additional genetic elements to ensure meiotic pairing and recombination. This effort requires innovation at multiple levels: designing MAC vectors encoding key functional elements, assembling large synthetic DNA constructs, and ultimately creating animals to test MACs in vivo. The proposed work builds on recent advances from the co-investigators’ teams in all of these areas, and we have key tools and expertise in place to build the necessary DNA templates, introduce them into embryos, analyze the outcomes, and develop alternative strategies as necessary. The most meaningful preliminary data would be to show a synthetic artificial chromosome that is successfully transmitted through mitosis and meiosis in vivo, but achieving this step is a major goal of our proposal and will require substantial investment of time and effort. Thus, we are requesting support for this project without the preliminary data that would demonstrate high likelihood of success, justifying consideration of our proposal as part of the T-R01 mechanism. We use mouse as a relatively rapid and tractable mammalian model system with outstanding opportunities for testing and debugging MACs, and our advances should readily transfer to other species for applications in biotechnology and medicine. Success in this project will represent a quantum leap in the development of synthetic artificial chromosome that are fully functional in vivo, providing unprecedented genome engineering capabilities in animal models and enabling diverse synthetic biology applications.

合成哺乳动物人工染色体（MAC）代表了基因组技术的新前沿， 具有改变染色体和合成生物学并刺激众多技术发展的潜力 医学的根本进步。人类基因组计划-Write旨在生成一整套合成人类 染色体。如果没有实现这一雄心勃勃的目标，MAC 在生物技术领域具有巨大的突破潜力 和医学，例如创建用于药物开发或收集患者的人性化动物模型 用于移植的个性化器官。此外，用最少的组件构建 MAC 将取得进展 我们对哺乳动物染色体的基本了解。 作为基因遗传的载体，功能齐全的染色体通过 有丝分裂和真核有性生殖和孟德尔遗传基础的特殊减数分裂 遗产。我们的目标是构建第一个通过种系实现忠实遗传的 MAC，使用 鼠标作为模型系统。其中一个障碍是着丝粒，即每条染色体上指导方向的位点。 通过有丝分裂和减数分裂传播。因为哺乳动物着丝粒不编码在 DNA 中 序列，目前尚不清楚如何构建包含这一关键元素的合成染色体。有 创造在减数分裂中作为同源染色体配对和重组的 MAC 还面临着额外的挑战。到 解决这些问题，我们将劫持现有的细胞机器来组装着丝粒染色质并 纳入额外的遗传元件以确保减数分裂配对和重组。 这项工作需要多个层面的创新：设计 MAC 向量编码关键功能 元素，组装大型合成 DNA 结构，并最终创造动物来测试体内 MAC。 拟议的工作建立在联合研究人员团队在所有这些领域的最新进展的基础上，我们 拥有关键工具和专业知识来构建必要的 DNA 模板，将其引入胚胎， 分析结果，并根据需要制定替代策略。最有意义的初步数据 将展示一种通过有丝分裂和减数分裂成功传递的合成人工染色体 体内，但实现这一步骤是我们提案的主要目标，需要大量时间投入 和努力。因此，我们请求对该项目的支持，但没有初步数据来证明 成功的可能性很高，证明我们的提案有理由考虑作为 T-R01 机制的一部分。 我们使用小鼠作为相对快速且易于处理的哺乳动物模型系统，具有出色的 测试和调试 MAC 的机会，我们的进步应该很容易转移到其他物种 在生物技术和医学中的应用。该项目的成功将代表着该领域的巨大飞跃 开发出在体内功能齐全的合成人工染色体，提供了前所未有的 动物模型中的基因组工程能力以及实现多种合成生物学应用。