Identifying N2-fixing microorganisms in photosynthetic microbial mats by Stable Isotope Probing (SIP) and nanometer-scale Secondary Ion Mass Spectrometry (nanoSIMS)

通过稳定同位素探测 (SIP) 和纳米级二次离子质谱 (nanoSIMS) 识别光合微生物垫中的固氮微生物

• 批准号: 164666922
• 负责人: Dr. Dagmar Woebken
• 金额: --
• 依托单位: Division of Microbial Ecology (DOME)
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2010-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/164666922?language=en
• 关键词: Identifying; N2; fixing; microorganisms; photosynthetic

Phototsynthetic microbial mats are important ecosystems in shallow-water marine environments since they supply these often nitrogen-limited environments with biologically available nitrogen, however the populations that fix nitrogen (diazotrophs) are poorly understood. Sequencing of the nifH genes and transcripts indicated that unicellular and filamentous cyanobacteria as well as heterotrophic bacteria could represent the dominant diazotrophs in microbial mats. However, the presence and/or transcription of this gene do not necessarily indicate that those organisms are primarily responsible for N2 fixation in this system due to transcriptional and posttranslational regulation. In the proposed research, the diazotrophic community of photosynthetic microbial mats will be studied using a combination of techniques. First, nifH gene and transcript sequencing will identify the potential diazotrophs and allows the design of primers and probes for the next task. Second, the abundance and seasonal succession of potential diazotrophs will be determined by nifH quantitative PCR and quantitative RT PCR. Third, this will be combined with 15N2 incubation experiments, in situ hybridization and investigations with nanometer-scale Secondary Ion Mass Spectrometry (nanoSIMS). In situ hybridization using halogen labeled tyramides and nanoSIMS (so called EL-FISH or HISH-SIMS) will assess the microbial community enriched in 15N, thereby allowing the identification of the microbial community that fixed N2 and the quantification of their metabolic activities at the single cell level.

光合微生物垫是浅水海洋环境中的重要生态系统，因为它们为这些通常氮有限的环境提供生物可利用的氮，但是固氮微生物（固氮生物）的种群却知之甚少。 nifH 基因和转录本的测序表明，单细胞和丝状蓝藻以及异养细菌可能代表微生物垫中的优势固氮菌。然而，该基因的存在和/或转录并不一定表明这些生物体由于转录和翻译后调节而主要负责该系统中的 N2 固定。在拟议的研究中，将使用多种技术的组合来研究光合微生物垫的固氮群落。首先，nifH 基因和转录本测序将鉴定潜在的固氮菌，并为下一步任务设计引物和探针。其次，潜在固氮生物的丰度和季节演替将通过 nifH 定量 PCR 和定量 RT PCR 确定。第三，这将与 15N2 孵化实验、原位杂交和纳米级二次离子质谱 (nanoSIMS) 研究相结合。使用卤素标记的酪酰胺和 nanoSIMS（所谓的 EL-FISH 或 HISH-SIMS）进行原位杂交将评估富含 15N 的微生物群落，从而能够识别固定 N2 的微生物群落并在单细胞水平上量化其代谢活动。