Analysis of Polyhydroxyalkanoates Granules in Haloferax mediterranei by Double-Fluorescence Staining with Nile Red and SYBR Green by Confocal Fluorescence Microscopy
| Autor: | Carmen Fernández-Ayuso, Lorena Simó-Cabrera, Carmen Pire, Fuensanta Monzó, Rosa María Martínez-Espinosa, Salvador Garcia-Chumillas, Verónica Cánovas |
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| Přispěvatelé: | Universidad de Alicante. Departamento de Agroquímica y Bioquímica, Universidad de Alicante. Instituto Multidisciplinar para el Estudio del Medio 'Ramón Margalef', Bioquímica Aplicada/Applied Biochemistry (AppBiochem) |
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0106 biological sciences
polyhydroxyalkanoates (PHAs) Polymers and Plastics Confocal Haloferax mediterranei Organic chemistry confocal microscopy 01 natural sciences Polyhydroxyalkanoates law.invention Polyhydroxyalkanoates (PHAs) 03 medical and health sciences chemistry.chemical_compound QD241-441 Confocal microscopy law 010608 biotechnology Fluorescence microscope SYBR Green haloarchaea 030304 developmental biology 0303 health sciences biology Chemistry Haloarchaea Nile red General Chemistry Bioquímica y Biología Molecular transmission electronic microscopy biology.organism_classification Staining Transmission electronic microscopy Biochemistry |
| Zdroj: | Polymers Volume 13 Issue 10 RUA. Repositorio Institucional de la Universidad de Alicante Universidad de Alicante (UA) Polymers, Vol 13, Iss 1582, p 1582 (2021) |
| ISSN: | 2073-4360 |
| DOI: | 10.3390/polym13101582 |
| Popis: | Haloferax mediterranei is a haloarchaeon of high interest in biotechnology because it produces and mobilizes intracellular polyhydroxyalkanoate (PHA) granules during growth under stress conditions (limitation of phosphorous in the culture media), among other interesting metabolites (enzymes, carotenoids, etc.). The capability of PHA production by microbes can be monitored with the use of staining-based methods. However, the staining of haloarchaea cells is a challenging task firstly, due to the high ionic strength of the medium, which is inappropriate for most of dyes, and secondly, due to the low permeability of the haloarchaea S-layer to macromolecules. In this work, Haloferax mediterranei is used as a halophilic archaeon model to describe an optimized protocol for the visualization and analysis of intracellular PHA granules in living cells. The method is based on double-fluorescence staining using Nile red and SYBR Green by confocal fluorescence microscopy. Thanks to this method, the capability of PHA production by new haloarchaea isolates could be easily monitored. |
| Databáze: | OpenAIRE |
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