Journal
BRAZILIAN JOURNAL OF MEDICAL AND BIOLOGICAL RESEARCH
Volume 50, Issue 1, Pages -Publisher
ASSOC BRAS DIVULG CIENTIFICA
DOI: 10.1590/1414-431X20165492
Keywords
Flow cytometry; Nile red; Poly-3-hydroxybutyrate; Herbaspirillum seropedicae; Azospirillum brasilense
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Funding
- National Institute of Science and Technology of Nitrogen Fixation INCT/CNPq/MCT
- CAPES
- PNPD/CAPES
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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 (R-2 = 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.
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