Mendelian inheritance of artificial chromosomes

人工染色体的孟德尔遗传

• 批准号: 10487450
• 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/10487450
• 关键词: Address; Animal Model; Animals; Area; Artificial Chromosomes; Artificial Mammalian Chromosomes; Behavior; Biological Models; Biotechnology; Cell division; Centromere; Chromatin; 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; Gold; 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; Problem Solving; 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.

人工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的潜力， 医学的根本进步。人类基因组计划旨在产生一整套合成人类基因组。 染色体如果没有这一雄心勃勃的目标，MAC在生物技术方面具有巨大的突破潜力 和医学，如创建用于药物开发的人源化动物模型或用于获取患者- 个性化器官移植此外，从最小组件构建MAC将取得进展 我们对哺乳动物染色体的基本理解。 作为遗传遗传的载体，功能齐全的染色体忠实地通过 有丝分裂和特殊的减数分裂分裂是真核生物有性生殖和孟德尔遗传的基础。 传承我们的目标是构建第一个通过种系实现忠实继承的MAC，使用 以老鼠为模型。一个障碍是着丝粒，即每条染色体上的基因座， 通过有丝分裂和减数分裂传播。因为哺乳动物的着丝粒并不是由DNA编码的 然而，目前还不清楚如何构建含有这一关键元素的合成染色体。有 创造在减数分裂中配对和重组为同源染色体的MAC的额外挑战。到 为了解决这些问题，我们将劫持现有的细胞机器来组装着丝粒染色质， 整合额外的遗传元件以确保减数分裂配对和重组。 这项工作需要在多个层面上进行创新：设计MAC矢量编码的关键功能 元件，组装大型合成DNA构建体，并最终创造动物来体内测试MAC。 拟议的工作建立在共同研究者团队在所有这些领域的最新进展的基础上，我们 拥有关键的工具和专业知识来构建必要的DNA模板，将它们引入胚胎， 分析结果，并在必要时制定替代战略。最有意义的初步数据 将展示一个人工合成的染色体，它能成功地通过有丝分裂和减数分裂传递 但实现这一步骤是我们建议的主要目标，需要投入大量时间 和努力因此，我们请求支持这个项目，但没有初步数据， 成功的可能性很高，因此有理由考虑将我们的建议作为T-R 01机制的一部分。 我们使用小鼠作为一个相对快速和易处理的哺乳动物模型系统， 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。这一项目的成功将标志着 人工合成染色体的发展，是完全功能的体内，提供前所未有的 在动物模型中的基因组工程能力，并使多样化的合成生物学应用。