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A lipophilic fluorescent LipidGreen1-based quantification method for high-throughput screening analysis of intracellular poly-3-hydroxybutyrate.

Abstract：

:Poly-3-hydroxybutyrate (PHB), the most abundant type of polyhydroxyalkanoates (PHA) is synthesized inside a variety of microorganisms as a primary candidate for industrial PHB production. Lipophilic dyes such as Nile red and BODIPY have been used to quantify intracellular PHB, but their uses have often been limited in terms of sensitivity and accuracy. In this study, a newly developed lipophilic fluorescent dye LipidGreen1 was used to quantify intracellular PHB. LipidGreen1 stained viable colonies by adding the dye into the medium which enabled the effective selection of PHB-positive cells. Furthermore, the fluorescence intensity of LipidGreen1 maintained its fluorescence intensity much longer than that of Nile red. The fluorescence intensities of intracellular PHB stained by LipidGreen1 accurately agreed with PHB contents measured by gas chromatography. In addition, internalization of LipidGreen1 in Escherichia coli cell was not necessary to obtain quantitative measurements. PHB-synthase mutants were differentiated by fluorescence intensities with a good correlation to increased levels of PHB production. These results show that LipidGreen1 is sensitive and accurate in high-throughput screening of newly isolated and genetically modified bacteria with enhanced PHB production.

journal_name

AMB Express

journal_title

AMB Express

authors

Choi JE,Na HY,Yang TH,Rhee SK,Song JK

doi

10.1186/s13568-015-0131-6

subject

Has Abstract

pub_date

2015-12-01 00:00:00

pages

131

issue

1

issn

2191-0855

journal_volume

5

pub_type

杂志文章

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