Please use this identifier to cite or link to this item: http://hdl.handle.net/10553/25087
Title: Using click-chemistry for visualizing in situ changes of translational activity in planktonic marine bacteria
Authors: Leizeaga, Ainara
Estrany, Margarita
Forn, Irene
Sebastián Caumel, Marta 
UNESCO Clasification: 251002 Oceanografía química
25 Ciencias de la tierra y del espacio
24 Ciencias de la vida
2510 Oceanografía
2414 Microbiología
Keywords: BONCAT
Marine bacteria
Click-chemistry
Marine ecology
Single-cell microbiology
Issue Date: 2017
Journal: Frontiers in Microbiology 
Abstract: A major challenge in microbial ecology is linking diversity and function to determine which microbes are actively contributing to processes occurring in situ. Bioorthogonal non-canonical amino acid tagging (BONCAT) is a promising technique for detecting and quantifying translationally active bacteria in the environment. This technique consists of incubating a bacterial sample with an analog of methionine and using click-chemistry to identify the cells that have incorporated the substrate. Here, we established an optimized protocol for the visualization of protein-synthesizing cells in oligotrophic waters that can be coupled with taxonomic identification using Catalyzed Reporter Deposition Fluorescent in Situ Hybridization. We also evaluated the use of this technique to track shifts in translational activity by comparing it with leucine incorporation, and used it to monitor temporal changes in both cultures and natural samples. Finally, we determined the optimal concentration and incubation time for substrate incorporation during BONCAT incubations at an oligotrophic site. Our results demonstrate that BONCAT is a fast and powerful semi-quantitative approach to explore the physiological status of marine bacteria.
URI: http://hdl.handle.net/10553/25087
ISSN: 1664-302X
DOI: 10.3389/fmicb.2017.02360
Source: Frontiers in Microbiology [ISSN 1664-302X], v. 8, article number 2360
Rights: by-nc-nd
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