scholarly journals Dataset of Nile Red Fluorescence Readings with Different Yeast Strains, Solvents, and Incubation Times

Data ◽  
2020 ◽  
Vol 5 (3) ◽  
pp. 77
Author(s):  
Mauricio Ramirez-Castrillon ◽  
Victoria Jaramillo-Garcia ◽  
Helio Barros ◽  
João Henriques ◽  
Valter Stefani ◽  
...  
Keyword(s):  
Culture Medium ◽  
Isopropyl Alcohol ◽  
Oleaginous Yeast ◽  
Nile Red ◽  
Solvent System ◽  
Oleaginous Yeasts ◽  
Yeast Strains ◽  
Nile Red Fluorescence ◽  
Accumulation Kinetics ◽  
Red Dye

We used Nile red to estimate lipid content in oleaginous yeasts using a high-throughput approach. We measured the fluorescence intensity of Nile red using different solvents, yeast strains, and incubation times in optimized excitation/emission wavelengths. The data show the relative fluorescence units (RFU) for Nile red excitation, using 1× PBS, 1× PBS and 5% v/v isopropyl alcohol, 50% v/v glycerol, culture medium A-gly broth, and A-gly broth supplemented with 5% v/v DMSO. In addition, we showed the RFU for the Nile red dye for different oleaginous and non-oleaginous yeast strains, such as Meyerozyma guilliermondii BI281A, Yarrowia lipolytica QU21 and Saccharomyces cerevisiae MRC164. Other measurements of lipid accumulation kinetics were shown for the above and additional yeast strains. These datasets provide the guidelines to obtain the optimal solvent system and the minimal interaction time for the Nile red dye to enter in the cells and obtain a stable readout.

scholarly journals Nile Red Incubation Time Before Reading Fluorescence Greatly Influences the Yeast Neutral Lipids Quantification

2021 ◽  
Vol 12 ◽  
Author(s):  
Mauricio Ramírez-Castrillón ◽  
Victoria P. Jaramillo-Garcia ◽  
Helio Lopes Barros ◽  
João A. Pegas Henriques ◽  
Valter Stefani ◽  
...  
Keyword(s):  
Lipid Content ◽  
High Throughput Screening ◽  
Oleaginous Yeast ◽  
Neutral Lipids ◽  
Nile Red ◽  
Oleaginous Yeasts ◽  
Red Fluorescence ◽  
Fluorescence Measurements ◽  
Yeast Strains ◽  
Nile Red Fluorescence

High-throughput screening methodologies to estimate lipid content in oleaginous yeasts use Nile red fluorescence in a given solvent and optimized excitation/emission wavelengths. However, Nile red fluorescence stabilization has been poorly analyzed, and high variability occurs when relative fluorescence is measured immediately or a few minutes after dye addition. The aim of this work was to analyze the fluorescence of Nile red at different incubation times using a variety of solvents and oleaginous/non-oleaginous yeast strains. We showed that fluorescence stabilization occurs between 20 and 30 min, depending on the strain and solvent. Therefore, we suggest that fluorescence measurements should be followed until stabilization, where Relative Fluorescence Units should be considered after stabilization for lipid content estimation.

scholarly journals Nile Red Staining for Oil Determination in Microalgal Cells: A New Insight through Statistical Modelling

2015 ◽  
Vol 2015 ◽  
pp. 1-14 ◽  
Author(s):  
Ronald Halim ◽  
Paul A. Webley
Keyword(s):  
Fluorescence Intensity ◽  
Nile Red ◽  
Biofuel Production ◽  
Staining Procedure ◽  
Surface Model ◽  
Second Phase ◽  
Red Fluorescence ◽  
Nile Red Fluorescence ◽  
Red Dye ◽  
Nile Red Staining

In the wake of global warming and rapid fossil fuel depletion, microalgae emerge as promising feedstocks for sustainable biofuel production. Nile red staining acts as a rapid diagnostic tool to measure the amount of biodiesel-convertible lipid that the cells accumulate. There is a need for the development of a more uniform staining procedure. In its first phase, this study examined the dependence of microalgal Nile red fluorescence (Tetraselmis suecica) in terms of its most pertinent staining variables. A quadratic surface model that successfully described the Nile red fluorescence intensity as a composite function of its variables was generated (r2=0.86). Cell concentration was shown to have a significant effect on the fluorescence intensity. Up to a certain threshold, fluorescence intensity was shown to increase with Nile red dye concentration. In its second phase, the study reviewed findings from previous Nile red studies to elucidate some of the fundamental mechanism underlying the diffusion of Nile red dye molecules into the microalgal cells and their subsequent interaction with intracellular lipids. Through the review process, we were able to develop a simple framework that provided a set of guidelines for the standardization of the Nile red staining procedure across different microalgal species.

scholarly journals Modified high-throughput Nile red fluorescence assay for the rapid screening of oleaginous yeasts using acetic acid as carbon source

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Catarina Miranda ◽  
Sara Bettencourt ◽  
Tatiana Pozdniakova ◽  
Joana Pereira ◽  
Paula Sampaio ◽  
...  
Keyword(s):  
Acetic Acid ◽  
Carbon Source ◽  
High Throughput ◽  
Nile Red ◽  
Fluorescence Assay ◽  
Rapid Screening ◽  
Oleaginous Yeasts ◽  
Red Fluorescence ◽  
Nile Red Fluorescence

scholarly journals Polyhydroxyalkanoates production by a bacterium isolated from mangrove soil samples collected from Quang Ninh province

2012 ◽  
Vol 3 (2) ◽  
pp. 76-79 ◽  
Author(s):  
Thuoc Van Doan ◽  
Binh Thi Nguyen
Keyword(s):  
Electron Microscope ◽  
Transmission Electron Microscope ◽  
Culture Medium ◽  
Carbon Sources ◽  
Nile Red ◽  
Soil Samples ◽  
Mangrove Soil ◽  
Transmission Electron ◽  
Bacterium Strain ◽  
Quang Ninh

A PHA producing bacterium (strain QN271) was selected from mangrove soil samples collected from Quang Ninh province by using the Nile red dying technique. PHA accumulation in the selected bacterium strain was confirmed by transmission electron microscope. With the exception of maltose or sucrose, the bacterium strain was found to be able to synthesize PHA from various carbon sources (glucose, xylose, fructose, glycerol, and glucose plus propionate). The strain accumulated poly(3-hydroxybutyrate) from glucose, fructose, xylose, and glycerol whereas poly(3-hydroxybutyrate-co-3-hydroxyvalarate) was produced when a combination of glucose and propionate was included in the culture medium. Fructose was found to be most suitable substrate for PHA synthesis by strain QN271. PHA content of 63.3% and CDW of 6 g/L were obtained after 32 hrs of cultivation in fructose medium. Chủng vi khuẩn có khả năng sinh tổng hợp PHA đã được phân lập từ đất rừng ngập mặn tỉnh Quảng Ninh nhờ kỹ thuật nhuộm với Nile red. Ảnh quan sát dưới kính hiển vi điện tử dẫn truyền chứng tỏ rằng chủng vi khuẩn này có khả năng tích lũy lượng lớn PHA trong tế bào. Chủng vi khuẩn tuyển chọn có khả năng sinh tổng hợp PHA từ nhiều nguồn các bon khác nhau như glucose, xylose, fructose, glucerol, glucose và propionate nhưng không có khả năng tổng hợp PHA từ maltose hoặc saccharose. Chủng vi khuẩn tuyển chọn tổng hợp poly (3-hydroxybutyrate) từ các nguồn các-bon như glucose, xylose, fructose, hay glycerol, trong khi đó poly (3-hydroxybutyrate-co-3-hydroxyvalarate) sẽ được tổng hợp khi phối hợp sử dụng hai nguồn các-bon (glucose và propionate). Fructose là nguồn các-bon tốt nhất cho chủng QN271 sinh tổng hợp PHA, khi nuôi cấy trong môi trường có fructose chủng vi khuẩn này có thể tạo ra lượng sinh khối là 6 g/L trong đó có chứa 63.3% PHA sau 32 giờ.

scholarly journals Generation of microplastics from the opening and closing of disposable plastic water bottles

2021 ◽  
Author(s):  
Tvisha Singh
Keyword(s):  
Particle Density ◽  
Nile Red ◽  
Bottled Water ◽  
Particle Generation ◽  
Large Spread ◽  
Total Particle ◽  
Contamination Levels ◽  
Opening And Closing ◽  
Red Dye ◽  
Disposable Plastic

Abstract There has recently been a significant increase in interest regarding the prevalence of microplastics in bottled water. Previous studies have shown that the composition of many of the microplastics in bottled water is consistent with the materials of the bottle and bottle cap. The focus of this study is to quantify microplastic particle generation from the cap and bottle interaction during open and close cycles. Nile Red dye was used for the detection of microplastics >4.7 μm in size. Microplastic contamination levels in the water were found to increase as the bottle cap is opened and closed repeatedly. The rate of generation of particles with bottle opening and closing cycles (553 ± 202 microplastics/L/cycle) is adequate to account for the total particle density in the water. This clearly demonstrates that the abrasion between the bottle cap and bottleneck is the dominant mechanism for the generation of microplastic contamination detected in bottled water. A large spread between the maximum and minimum levels of microplastic contamination for bottles from the same lot, regardless of the number of times the cap is opened and closed, suggests that mechanical tolerances in the manufacturing of bottles and caps might play an important role in microplastic generation.

scholarly journals Engineering the Oleaginous Yeast Rhodosporidium toruloides for Improved Resistance Against Inhibitors in Biomass Hydrolysates

2021 ◽  
Vol 9 ◽  
Author(s):  
Liting Lyu ◽  
Yadong Chu ◽  
Sufang Zhang ◽  
Yue Zhang ◽  
Qitian Huang ◽  
...  
Keyword(s):  
Lignocellulosic Biomass ◽  
Oleaginous Yeast ◽  
Lipid Production ◽  
Cell Mass ◽  
Peroxidase Gene ◽  
The Past ◽  
Yeast Strains ◽  
Biomass Hydrolysates ◽  
Improved Performance ◽  
Inhibitor Resistance

Conversion of lignocellulosic biomass into lipids and related chemicals has attracted much attention in the past two decades, and the oleaginous yeast Rhodosporidiumtoruloides has been widely used in this area. While R. toruloides species naturally have physiological advantages in terms of substrate utilization, lipid accumulation, and inhibitor resistance, reduced lipid production and cell growth are noticed when biomass hydrolysates are used as feedstocks. To improve the robustness of R. toruloides, here, we devised engineered strains by overexpressing genes responsible for phenolic compound degradation. Specifically, gene expression cassettes of the manganese peroxidase gene (MNP) and versatile peroxidase gene (VP) were constructed and integrated into the genome of R. toruloides NP11. A series of engineered strains were evaluated for lipid production in the presence of typical phenolic inhibitors. The results showed that R. toruloides strains with proper expression of MNP or VP indeed grew faster in the presence of vanillin and 5-hydroxymethylfurfural than the parental strain. When cultivated in concentrated mode biomass hydrolysates, the strain VP18 had improved performance as the cell mass and lipid content increased by 30% and 25%, respectively. This study provides more robust oleaginous yeast strains for microbial lipid production from lignocellulosic biomass, and similar efforts may be used to devise more advanced lipid producers.

scholarly journals Silicon Nanomembrane Filtration and Imaging for the Evaluation of Microplastic Entrainment along a Municipal Water Delivery Route

2020 ◽  
Vol 12 (24) ◽  
pp. 10655
Author(s):  
Gregory R. Madejski ◽  
S. Danial Ahmad ◽  
Jonathan Musgrave ◽  
Jonathan Flax ◽  
Joseph G. Madejski ◽  
...  
Keyword(s):  
Drinking Water ◽  
Water Transport ◽  
Transport Process ◽  
Nile Red ◽  
Machine Learning Algorithms ◽  
Hydrophobic Polymers ◽  
Silicon Nanomembrane ◽  
Debris Transport ◽  
End Stage ◽  
Red Dye

To better understand the origin of microplastics in municipal drinking water, we evaluated 50 mL water samples from different stages of the City of Rochester’s drinking water production and transport route, from Hemlock Lake to the University of Rochester. We directly filtered samples using silicon nitride nanomembrane filters with precisely patterned slit-shaped pores, capturing many of the smallest particulates (<20 µm) that could be absorbed by the human body. We employed machine learning algorithms to quantify the shapes and quantity of debris at different stages of the water transport process, while automatically segregating out fibrous structures from particulate. Particulate concentrations ranged from 13 to 720 particles/mL at different stages of the water transport process and fibrous pollution ranged from 0.4 to 8.3 fibers/mL. A subset of the debris (0.2–8.6%) stained positively with Nile red dye which identifies them as hydrophobic polymers. Further spectroscopic analysis also indicated the presence of many non-plastic particulates, including rust, silicates, and calcium scale. While water leaving the Hemlock Lake facility is mostly devoid of debris, transport through many miles of piping results in the entrainment of a significant amount of debris, including plastics, although in-route reservoirs and end-stage filtration serve to reduce these concentrations.

161 LIPID CONTENT, RESISTANCE TO CRYOPRESERVATION, AND SEX RATIO OF IN VITRO BOVINE BLASTOCYSTS PRODUCED IN A SERUM-FREE SYSTEM

2006 ◽  
Vol 18 (2) ◽  
pp. 188
Author(s):  
F. George ◽  
C. Daniaux ◽  
G. Genicot ◽  
F. Focant ◽  
B. Verhaeghe ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Lipid Content ◽  
Culture Medium ◽  
Nile Red ◽  
Embryo Cryopreservation ◽  
Hatching Rate ◽  
Free System ◽  
Serum Free

In vitro-produced (IVP) bovine blastocysts are known to be more sensitive to cryopreservation than their in vivo counterparts. Removing serum from the culture medium decreases sanitary risk and could improve embryo resistance to cryopreservation by preventing the accumulation of intracellular lipids. Our objectives were to evaluate the lipid content, resistance to cryopreservation, and sex ratio of IVP embryos cultured in a serum-free system. Oocytes from slaughterhouse ovaries were matured in a serum-free enriched medium (Donnay et al. 2004 Reprod. Fertil. Dev. 16, 274) and cultured in 5% O2 in modified SOF supplemented with 5% FCS (FCS) or with insulin-transferrin-selenium (ITS) and 0.1 mg/mL polyvinylpyrrolidone (PVP) (ITS-PVP) or 4 mg/mL BSA (ITS-BSA) (Daniaux et al. 2005 Reprod. Fertil. Dev. 17, 217). Day 5 morulae were stained with the fluorescent dye Nile Red in order to evaluate their lipid content (Genicot et al. 2005 Theriogenology 63, 1181). Day 7 blastocysts (diameter ≥160 µm) were selected, classified according to their size, and frozen in HEPES-SOF containing 1.5 M ethylene glycol, 0.1 M sucrose, and 1.8 mg/mL wheat peptones (George et al. 2002 Reproduction 29, 51). The lipid content was significantly lower in morulae cultured in ITS-BSA compared with the two other media (320 ± 10 arbitrary fluorescence units vs. 383 ± 12 in FCS and 406 ± 10 in ITS-PVP; n = 271; ANOVA2: P < 0.01). After cryopreservation, a higher total hatching rate was found 24 h post-thawing in blastocysts cultured in ITS-BSA and for both serum-free conditions at 48 h (Table 1). In particular, embryos ≤180 µm cultured in FCS were less resistant to cryopreservation than embryos of the same size produced without serum. Expanded blastocysts cultured in ITS-BSA were sexed by PCR (Grisart et al. 1995 Theriogenology 43, 1097) and a higher proportion of male embryos was found (62.7%; n = 51). In conclusion, a complete serum-free system was set up from oocyte maturation to embryo cryopreservation that gave high quality embryos resistant to cryo-preservation. Embryos produced in ITS-BSA presented a lower lipid content, but a shift of the expanded blastocyst sex ratio toward males was observed. Table 1. Hatching rates post-thawing as a function of the blastocyst size and the culture medium

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