A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry
| Autor: | L.P.S. Alves, A.T. Almeida, L.M. Cruz, F.O. Pedrosa, E.M. de Souza, L.S. Chubatsu, M. Müller-Santos, G. Valdameri |
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| Jazyk: | angličtina |
| Předmět: | |
| Zdroj: | Brazilian Journal of Medical and Biological Research, Vol 50, Iss 1 |
| Druh dokumentu: | article |
| ISSN: | 1414-431X 1414-431x |
| DOI: | 10.1590/1414-431x20165492 |
| Popis: | The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots. |
| Databáze: | Directory of Open Access Journals |
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