IOS EDGE: Development of genetic tools for the dominant phototroph in the sea

IOS EDGE：开发海洋中优势光养生物的遗传工具

• 批准号: 1645061
• 负责人: Sallie Chisholm
• 金额: $ 89万
• 依托单位: Massachusetts Institute of Technology
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1645061&HistoricalAwards=false
• 关键词: IOS; EDGE; Development; genetic; tools

Phytoplankton are microscopic plants that form the base of ocean food chains and collectively do half of the photosynthesis on Earth, equal to all the plants on land. Prochlorococcus is the single most abundant phytoplankton species in the global oceans and is responsible for producing an estimated 4 billion tons of fixed carbon each year, which is approximately the same productivity as global croplands. In addition they are the smallest and most efficient photosynthesizers designed by nature, relying only on sunlight, CO2 and essential nutrients for growth. For the past 25 years, research on this minimal prototroph has led to a growing understanding of its features across all scales of biological organization, from the genome to the global ecosystem. This fascinating microbe has also offered a great opportunity to foster public interest on topics such as ocean ecology, biodiversity and genomics. It attracts the attention of science journalists, and serves as an "ambassador" for microorganisms - exemplifying their important role in planetary maintenance. The one thing that is missing from the Prochlorococcus model system tool kit is the ability to do genetics. This bacterium has resisted all genetic manipulation attempts to date, which has significantly hampered the rate of discovery. The goal of this project is to directly address this limitation by developing an efficient genetic system for Prochlorococcus using both traditional and state-of-the-art tools developed specifically for this project.Research on Prochlorococcus has expanded steadily over the past 25 years, as more and more strains have been brought into culture, and their distribution in the global oceans has been mapped extensively. The PIs now have 100s of Prochlorococcus cultures in the lab, an extensive amount of transcriptomics and proteomics data, and they continue to accumulate genomic data from cultures and wild populations. However each new Prochlorococcus strain, or single cell sequenced, adds roughly 100 entirely new genes to the Prochlorococcus pangenome, the vast majority of which are of unknown function. The development of a genetic system is necessary for identifying the functions of these unknown genes and testing hypotheses involving those with putative functions. This project notably involves a high throughput microfluidic electroporation technology to transform recalcitrant microbes orders of magnitude faster than traditional approaches. A global mutagenesis (Transposon-based tools) and targeted mutagenesis (CRISPR-based tools) strategy will be implemented for Prochlorococcus, while optimizing the microfluidic electroporation technology to its full potential. As proof-of-concept, each tool will be tested in its ability to efficiently reveal gene functions by using our current known repertoire of nutrient acquisition genes in Prochlorococcus.

浮游植物是构成海洋食物链基础的微型植物，它们共同完成地球上一半的光合作用，相当于陆地上所有的植物。 原绿球藻是全球海洋中最丰富的浮游植物物种，每年产生约40亿吨固定碳，这与全球农田的生产力大致相同。此外，它们是自然界设计的最小和最有效的光合作用器，仅依赖于阳光，二氧化碳和生长所必需的营养物质。在过去的25年里，对这种最小原养型的研究使人们越来越多地了解它在所有生物组织尺度上的特征，从基因组到全球生态系统。这种迷人的微生物也提供了一个很好的机会，以促进公众对海洋生态学，生物多样性和基因组学等主题的兴趣。它吸引了科学记者的注意力，并作为微生物的“大使”-证明了它们在行星维护中的重要作用。原绿球藻模型系统工具包缺少的一件事是遗传学的能力。这种细菌抵抗了迄今为止所有的基因操作尝试，这大大阻碍了发现的速度。该项目的目标是直接解决这一限制，通过开发一个有效的遗传系统，为原绿球藻使用传统的和国家的最先进的工具，专门为此项目开发。在过去的25年里，对原绿球藻的研究稳步扩大，越来越多的菌株已被带入文化，他们在全球海洋中的分布已被广泛绘制。PI现在在实验室中有100个原绿球藻培养物，大量的转录组学和蛋白质组学数据，他们继续积累来自培养物和野生种群的基因组数据。然而，每一个新的原绿球藻菌株，或测序的单细胞，都为原绿球藻pangenome增加了大约100个全新的基因，其中绝大多数是未知功能的。遗传系统的发展是必要的，以确定这些未知基因的功能和测试假设涉及那些假定的功能。该项目特别涉及高通量微流体电穿孔技术，以比传统方法快几个数量级的速度转化寄生微生物。将对原绿球藻实施全局诱变（基于转座子的工具）和靶向诱变（基于CRISPR的工具）策略，同时优化微流体电穿孔技术以充分发挥其潜力。作为概念验证，每个工具都将通过使用我们目前已知的原绿球藻营养获取基因库来测试其有效揭示基因功能的能力。

项目成果

Association of Lanthipeptide Genes with TnpAREP Transposases in Marine Picocyanobacteria

海洋微微蓝藻中 Lanthipeptide 基因与 TnpAREP 转座酶的关联

• DOI: 10.1101/2020.03.09.984088
• 发表时间: 2020
• 期刊: bioRxiv
• 作者: Laurenceau, R.;Raho, N;Cariani, Z;Bliem, C;Osman, M;Chisholm, SW
• 通讯作者: Chisholm, SW

Toward a genetic system in the marine cyanobacterium Prochlorococcus.

迈向海洋蓝细菌核酸菌群中的遗传系统。

• DOI: 10.1099/acmi.0.000107
• 发表时间: 2020
• 期刊: Access microbiology
• 作者: Laurenceau R;Bliem C;Osburne MS;Becker JW;Biller SJ;Cubillos-Ruiz A;Chisholm SW
• 通讯作者: Chisholm SW

Novel integrative elements and genomic plasticity in ocean ecosystems

• DOI: 10.1016/j.cell.2022.12.006
• 发表时间: 2023-01-05
• 期刊: CELL
• 影响因子: 64.5
• 作者: Hackl，Thomas;Laurenceau，Raphael;Chisholm，Sallie W.
• 通讯作者: Chisholm，Sallie W.