表型驱动的单细胞基因组技术研究海洋微型蓝细菌的生物多样性及其环境适应机制

Marine picocyanobacteria，including genera Prochlorococcus and Synechococcus, numerically dominate the picophytoplankton in the world ocean, and make an important contribution to global primary production. Picocyanobacteria have evolved into diverse ecotypes to adapt to different marine habitats. Genomic analyses provide the best interpretation for their widespread distribution. Metagenomics has been widely used to study the microbial community structure and metabolic potential, however it shows the average properties of community and ignores individual heterogeneity. As an important complement technology to metagenomics, single-cell genomics can reveal the high heterogeneity of marine microbes at the individual level, especially for the novel phylogenetic or functional groups. .In this project, we use picocyanobacteria as studied subjects and develop a phenotype-driven single-cell genomics method to investigate their biodiversity and survival strategies. .1) A multi-laser flow cytometry method could determine the relative fluorescence action spectra of picocyanobacteria at the single-cell and population-level. By coupling the fluorescence spectra to statistical analysis, we would like to establish a rapid phenotypic analysis technology for picocyanobacteria..2) Phenotype-driven single-cell genomics provides a solution by using a phenotype-based screen to subsample complex microbial communities in a targeted manner for the isolation and genome sequencing of single cells. It’s helpful for the discovery of novel phylogenetic or functional clades in picocyanobacteria..3) Comparative genomics among close-related species could improve our understanding to the vertical and horizontal evolutionary processes of specific genes among picocyanobacteria. .4）In this project, we put the picocyanobacterial genomics into an environmental context and develop a comprehensive analysis of the environment-phenotype-genotype paradigm to give insights into the adaptations of these marine microbes to local environment and how this is reflected at the genomic level..Through this project, it can help us to understand the biodiversity of picocyanobacteria, and provides new insights into their biogeochemistry contribution in global ocean. It is critical for understanding and modeling the global climate change using obtained information from studied marine cyanobacteria model.

蓝细菌是海洋初级生产力的主要贡献者，在缓解全球气候变化中扮演着重要重要。蓝细菌在全球海洋有着广泛分布，形成适应不同海洋环境的生态类型。基因组学研究为蓝细菌的全球分布提供了最好的诠释。然而，宏基因组学揭示的是群落结构的平均属性，忽视个体异质性。本项目利用表型驱动的单细胞基因组技术，基于流式细胞分析分选平台，建立蓝细菌表型特征分析技术；通过对特殊表型的二次分选，提高单细胞基因组的效率，揭示其高度异质性，并有助于新系统发育类群或功能类群的发现。通过对不同亲缘关系蓝细菌基因组的比较分析，揭示其垂直进化和水平演化过程；并将基因组信息融入环境背景，结合表型特征分析，建立“环境-表型-基因”的有机结合，深入解析蓝细菌的遗传多样性、系统发育、生态位分化在基因组水平上的响应。本项目将为更好理解蓝细菌对于全球气候变化的响应提供重要见解，为蓝细菌的开发利用提供科学依据，对于应对全球气候变化具有重要科学价值。

蓝细菌在全球海洋有着广泛分布，是海洋初级生产力的主要贡献者。项目组利用流式细胞技术，基于细胞大小、色素组成等表型特征，针对性地对蓝细菌进行分离纯化，结合单细胞基因组扩增技术，建立了海洋蓝细菌的微型宏基因组技术，并将其应用于厦门近岸海域聚球藻群落结构和功能基因组学的研究。研究结果表明，基于表型驱动的单细胞基因组扩增技术，不仅能够获得低丰度聚球藻的基因组信息，在更深层次上揭示蓝细菌的遗传多样性；而且在厦门海域的首次应用就发现了131种聚球藻新功能基因，将有助于深入理解聚球藻的遗传多样性与环境适应机制。.在原绿球藻生态型、基因组和环境适应机制方面，项目组在西太平洋和南海分离纯化了67株原绿球藻，并最终获得了15个原绿球藻全基因组草图序列，其中包括13个高光II型（HL-II）和2个低光I型（LL-I）。比较基因组学分析结果表明，原绿球藻存在复杂多样的亚生态型，其中亚生态型附属基因组的进化与核心基因组SNP的进化相互对应，共同推动了各亚类群的进化、分化和环境适应。宏基因组学和单细胞基因组学的结合，在研究原绿球藻亚生态型的生态分布、功能基因差异以及环境适应机制中将发挥重要作用。.在环境适应机制方面，项目组从蓝细菌与异养细菌的相互作用关系入手，通过宏基因组学手段对蓝细菌与异养细菌共培养体系进行分析。异养细菌不仅含有许多氮、磷相关的代谢基因，具有多种营养和能量代谢能力；而且包含多种肽酶、糖苷水解酶以及与黏附相关的基因，可能对于蓝细菌代谢产物的降解与转化具有重要意义。通过对蓝细菌及其共培养微生物的研究，将有助于深入理解海洋蓝细菌的生活方式及其代谢途径，为研究蓝细菌的环境适应机制提供不一样的视角，对于准确理解海洋生物地球化学循环具有重要意义。

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