Antibiotic Dereplication Using the Antibiotic Resistance Platform
| Autor: | Haley L. Zubyk, Georgina Cox, Gerard D. Wright |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
medicine.drug_class
General Chemical Engineering Antibiotics Drug resistance Secondary metabolite 010402 general chemistry medicine.disease_cause 01 natural sciences General Biochemistry Genetics and Molecular Biology Mass Spectrometry law.invention chemistry.chemical_compound Antibiotic resistance law Drug Discovery medicine Escherichia coli Biological Products Natural product Chromatography General Immunology and Microbiology 010405 organic chemistry General Neuroscience Petri dish Drug Resistance Microbial 0104 chemical sciences Anti-Bacterial Agents chemistry Fermentation medicine.drug |
| Zdroj: | Journal of visualized experiments : JoVE. (152) |
| ISSN: | 1940-087X |
| Popis: | One of the main challenges in the search for new antibiotics from natural product extracts is the re-discovery of common compounds. To address this challenge, dereplication, which is the process of identifying known compounds, is performed on samples of interest. Methods for dereplication such as analytical separation followed by mass spectrometry are time-consuming and resource-intensive. To improve the dereplication process, we have developed the antibiotic resistance platform (ARP). The ARP is a library of approximately 100 antibiotic resistance genes that have been individually cloned into Escherichia coli. This strain collection has many applications, including a cost-effective and facile method for antibiotic dereplication. The process involves the fermentation of antibiotic-producing microbes on the surface of rectangular Petri dishes containing solid medium, thereby allowing for the secretion and diffusion of secondary metabolites through the medium. After a 6 day fermentation period, the microbial biomass is removed, and a thin agar-overlay is added to the Petri dish to create a smooth surface and enable the growth of the E. coli indicator strains. Our collection of ARP strains is then pinned onto the surface of the antibiotic-containing Petri dish. The plate is next incubated overnight to allow for E. coli growth on the surface of the overlay. Only strains containing resistance to a specific antibiotic (or class) grow on this surface enabling rapid identification of the produced compound. This method has been successfully used for the identification of producers of known antibiotics and as a means to identify those producing novel compounds. |
| Databáze: | OpenAIRE |
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