人工基因组结构变异与碱基突变迭代进化策略的研究

Genomic structural variation and genomic base mutations are the main forms of biological evolution, and are expected to be used for the development of the desired phenotype of microorganisms. The key scientific problem that applicants intend to solve is how to quickly achieve genome reconstruction and expected functional evolution through genomic structural variation and base mutations. The aim of this study was to reveal the molecular mechanism of the evolution of genomic structure variation and base mutations to promote evolution, and to provide new techniques generating strain with desired phenotypes rapidly. The research contents mainly include: 1) Multi-scale evolutionary strategies of genomic DNA, designing and controlling the cooperative combination of artificial yeast genome rearrangement system and plasma mutagenesis system; 2) Iteratively evolving high-producing astaxanthin yeast with high yield of astaxanthin as the target Multiple iterations on the basis of collaborative binding strategies to increase evolutionary speed and expand library diversity; 3) Functional discovery of evolutionary genomes; Reveal the relationship between genome changes and function of desired strains through genome re-sequencing, etc. This study is expected to establish a new method for synergistic binding of genomic structural variation and base mutations, and to rapidly obtain astaxanthin-producing yeast strains, revealing related targets gene and mechanisms of astaxanthin synthesis.

基因组结构变异与基因组碱基突变是生物进化的主要形式，可望用作开发大幅度提高微生物预期功能性状的方法。申请者拟解决的关键科学问题是如何通过基因组结构变异与碱基突变快速实现基因组再造及预期功能进化。本研究旨在揭示基因组结构变异与碱基突变协同促进进化的分子机制，为快速获得优势性状菌株提供新技术。研究内容主要包括：1）基因组DNA多尺度进化策略，设计并控制人工酵母基因组重排系统与等离子体诱变系统的协同结合；2）迭代进化高产虾青素酵母，以高产虾青素为目标，在协同结合策略的基础上进行多轮迭代，提高进化速度并拓展文库多样性；3）进化基因组的功能发现；通过基因组重测序等揭示优势菌株的基因组变化与功能之间的关系，解析相关作用靶点。本研究预期建立基因组结构变异与碱基突变协同结合的新方法,快速获得高产虾青素酵母菌株，揭示虾青素合成相关靶点和机理。

理性改造能够有效提高人工细胞合成化学品的生产能力。但是当前对基因、代谢调控网络的认知水平还不够充分，限制了其对目标化学品(如虾青素)合成的高效改造。传统基因组诱变主要是基于DNA损伤修复，主要产生单碱基等小尺度的突变。人工基因组重排基于重组酶系统拓扑异构原理产生复制、删除、易位和倒置等大尺度的突变。本项目开展设计基因组多尺度进化新技术，提高虾青素生物合成以及进化基因组结构功能发现这三方面开展研究，完成了全部研究内容，达到了预期目标。拓展基因组重排系统的设计原则，建立基因组结构变异与碱基突变迭代方法，揭示了基因组序列排列方式与功能的内在联系；建立了虾青素外源模块体内体外重排方法，实现外源功能模块与底盘细胞的快速适配；建立跨染色体重排方法，揭示了跨染色体重排的变化规律和驱动表型进化的遗传机制；形成若干具有自主知识产权的合成生物技术，获得高产虾青素菌株生产效率高，具有应用前景。