The Experts below are selected from a list of 162 Experts worldwide ranked by ideXlab platform
Ruud A Weusthuis - One of the best experts on this subject based on the ideXlab platform.
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Stable transformation of the green algae Acutodesmus Obliquus and Neochloris oleoabundans based on E. coli conjugation
Algal Research-Biomass Biofuels and Bioproducts, 2019Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Sarah D'adamo, Ruud A WeusthuisAbstract:Abstract Microalgae are an ideal platform for the production of high-value chemicals, nutritional products and biofuels. Genetic engineering could speed up the development of microalgae derived products and reduce the overall production costs. Genetic methods such as particle bombardment, electroporation, Agrobacterium tumefaciens mediated transformation (ATMT), and agitation with glass beads and silicon carbide whiskers have been developed for the genetic transformation of microalgae. However, the transformation efficiency is species dependent, so a variety of transformation methods are required to engineer a wide range of microalgae species. The oleaginous microalgae Acutodesmus Obliquus and Neochloris oleoabundans have a great potential as production platforms due to their ability to produce large amounts of triacylglycerol (TAG). Genetic modification techniques however are required to increase TAG levels further or to modify the fatty acid composition. Recently, a conjugation-based method for the delivery of episomes from bacteria to diatom microalgae has been reported. In this study, we have achieved the successful transformation of green oleaginous microalgal strains by transferring an expression vector via conjugation from E. coli. Since delivery of exogenous DNA into the microalgae cells is only the first step in obtaining transgenic microalgae, we further analyzed transformation efficiencies by PCR and expression of the Clover fluorescent protein in the targeted species.
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improved dna protein delivery in microalgae a simple and reliable method for the prediction of optimal electroporation settings
Algal Research-Biomass Biofuels and Bioproducts, 2018Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Lenny De Jaeger, Ka Y F Lip, Justus W J Olijslager, Jan Springer, Emil J H Wolbert, Dirk E Martens, Gerrit Eggink, Ruud A WeusthuisAbstract:Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus Obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.
Camilo F. Muñoz - One of the best experts on this subject based on the ideXlab platform.
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Stable transformation of the green algae Acutodesmus Obliquus and Neochloris oleoabundans based on E. coli conjugation
Algal Research-Biomass Biofuels and Bioproducts, 2019Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Sarah D'adamo, Ruud A WeusthuisAbstract:Abstract Microalgae are an ideal platform for the production of high-value chemicals, nutritional products and biofuels. Genetic engineering could speed up the development of microalgae derived products and reduce the overall production costs. Genetic methods such as particle bombardment, electroporation, Agrobacterium tumefaciens mediated transformation (ATMT), and agitation with glass beads and silicon carbide whiskers have been developed for the genetic transformation of microalgae. However, the transformation efficiency is species dependent, so a variety of transformation methods are required to engineer a wide range of microalgae species. The oleaginous microalgae Acutodesmus Obliquus and Neochloris oleoabundans have a great potential as production platforms due to their ability to produce large amounts of triacylglycerol (TAG). Genetic modification techniques however are required to increase TAG levels further or to modify the fatty acid composition. Recently, a conjugation-based method for the delivery of episomes from bacteria to diatom microalgae has been reported. In this study, we have achieved the successful transformation of green oleaginous microalgal strains by transferring an expression vector via conjugation from E. coli. Since delivery of exogenous DNA into the microalgae cells is only the first step in obtaining transgenic microalgae, we further analyzed transformation efficiencies by PCR and expression of the Clover fluorescent protein in the targeted species.
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improved dna protein delivery in microalgae a simple and reliable method for the prediction of optimal electroporation settings
Algal Research-Biomass Biofuels and Bioproducts, 2018Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Lenny De Jaeger, Ka Y F Lip, Justus W J Olijslager, Jan Springer, Emil J H Wolbert, Dirk E Martens, Gerrit Eggink, Ruud A WeusthuisAbstract:Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus Obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.
Wook Jin Choi - One of the best experts on this subject based on the ideXlab platform.
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Effect of trophic conditions on microalga growth, nutrient removal, algal organic matter, and energy storage products in Scenedesmus (Acutodesmus) Obliquus KGE-17 cultivation
Bioprocess and Biosystems Engineering, 2019Co-Authors: Wook Jin Choi, Joonhong Park, Kyung Guen Song, A. Na Chae, Byung Chan LeeAbstract:This study compared the performance of microalga growth, nutrient removal, algal organic matter, and energy storage products in mixotrophic, photoautotrophic, and heterotrophic conditions. Scenedesmus Obliquus was used as a model species. Mixotrophic condition showed the highest specific growth rate of 0.96 d^−1 as well as the fastest nitrogen and phosphorus removal rate of 85.17 mg-N g-cell^−1 day^−1 and 11.49 mg-P g-cell^−1 day^−1, respectively, compared with photoautotrophic and heterotrophic conditions. Mixotrophic microalgae had relatively higher carbohydrates and lipids contents (21.8 and 24.0%) than photoautotrophic and heterotrophic conditions. Meanwhile, algal organic matter (AOM) in the medium was produced at the highest level under photoautotrophic condition. Mixotrophic condition was more efficient in terms of microalga growth, nutrient removal, production of energy storage products, and suppression of AOM, and would be adaptable for wastewater treatment process.
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treating high strength saline piggery wastewater using the heterotrophic cultivation of Acutodesmus Obliquus
Biochemical Engineering Journal, 2016Co-Authors: Wook Jin Choi, Na A Chae, Joonhong Park, Kyung Guen SongAbstract:Abstract This study combined an algae treatment with post-ozonation for the robust treatment of piggery effluent in which high levels of organic constituents, inorganic nutrients, color, and salts remained. Due to a nearly complete light limitation resulting from a high level of color, the algae treatment was conducted with continuous O 2 supplementation instead of using the combination of high lighting and CO 2 injection. The microalga Acutodesmus Obliquus KGE-17 showed tolerance to high salinity (up to 5.2% as chloride) and was capable of utilizing dissolved organics (1923 mg-COD/(g-cell)(day)) under the heterotrophic growth conditions. The use of A . Obliquus for remediation of piggery effluent also resulted in an operational simplicity in the simultaneous removal of nitrogen and phosphorus in a single treatment process, in contrast to conventional biological nutrient removal processes. Subsequent ozonation was successful in decolorizing the algae-treated wastewater and in improving the biodegradability of residual organics in post-treatment processes.
Muñoz, Camilo F. - One of the best experts on this subject based on the ideXlab platform.
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Stable transformation of the green algae Acutodesmus Obliquus and Neochloris oleoabundans based on E. coli conjugation
'Elsevier BV', 2019Co-Authors: Muñoz, Camilo F., Sturme, Mark H.j., D'adamo Sarah, Weusthuis, Ruud A., Wijffels, Rene HubertusAbstract:Microalgae are an ideal platform for the production of high-value chemicals, nutritional products and biofuels. Genetic engineering could speed up the development of microalgae derived products and reduce the overall production costs. Genetic methods such as particle bombardment, electroporation, Agrobacterium tumefaciens mediated transformation (ATMT), and agitation with glass beads and silicon carbide whiskers have been developed for the genetic transformation of microalgae. However, the transformation efficiency is species dependent, so a variety of transformation methods are required to engineer a wide range of microalgae species. The oleaginous microalgae Acutodesmus Obliquus and Neochloris oleoabundans have a great potential as production platforms due to their ability to produce large amounts of triacylglycerol (TAG). Genetic modification techniques however are required to increase TAG levels further or to modify the fatty acid composition. Recently, a conjugation-based method for the delivery of episomes from bacteria to diatom microalgae has been reported. In this study, we have achieved the successful transformation of green oleaginous microalgal strains by transferring an expression vector via conjugation from E. coli. Since delivery of exogenous DNA into the microalgae cells is only the first step in obtaining transgenic microalgae, we further analyzed transformation efficiencies by PCR and expression of the Clover fluorescent protein in the targeted species.publishedVersionPaid Open Acces
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Effect of Single and Combined Expression of Lysophosphatidic Acid Acyltransferase, Glycerol-3-Phosphate Acyltransferase, and Diacylglycerol Acyltransferase on Lipid Accumulation and Composition in Neochloris oleoabundans
2019Co-Authors: Muñoz, Camilo F., Weusthuis, Ruud A., D’adamo Sarah, Wijffels, René H.Abstract:Microalgal lipids are promising feedstocks for food and biofuels. Since lipid production by microalgae is not yet economically feasible, genetic engineering is becoming a promising strategy to achieve higher lipid accumulation and productivities. Enzymes involved in the Kennedy pathway such as glycerol-3-phosphate acyltransferase (GPAT), lysophosphatidic acid acyltransferase (LPAT), and diacylglycerol acyltransferase (DGAT) catalyze key steps in the formation of triacylglycerol, which is the main constituent of lipids in N. oleoabundans. The overexpression of these enzymes in the targeted strain has a great potential to further increase their triacylglycerol content. We overexpressed single and multiple encoding genes for LPAT, GPAT, and DGAT from Acutodesmus Obliquus in N. oleoabundans. Strains overexpressing single genes produced up to 52% and 45% g · gDW-1, which corresponds to 1.3- and 1.4-fold increase in total fatty acids and triacylglycerols, respectively. The orchestrated expression of the three genes resulted in 49% and 39% g · gDW-1, which is 1.2-folds increase in total fatty acids and triacylglycerols. Single expression of LPAT, GPAT, and DGAT genes resulted in higher lipid productivities during starvation without a significant effect on growth and photosynthetic activity during replete conditions. On the other hand, the simultaneous expression of LPAT, GPAT, and DGAT genes resulted in 52% lower growth rate, 14% lower photosynthetic activity and 4-folds increase in cell diameter. Moreover, the multigene expressing line showed a decrease in carbohydrates and protein content and an increase in pigments during nitrogen starved condition. The single and multiple expression of heterologous genes LPAT, GPAT, and DGAT showed to significantly enhanced the lipid accumulation in N. oleoabundans. Single gene expression resulted in higher lipid production and productivities without having a significant impact in the physiological status of the strains. This approach shows the potential for the generation of microalgal strains with higher economical potential for the production of lipids.
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Effect of single and combined expression of Lysophosphatidic Acid Acyltransferase, Glycerol-3-Phosphate Acyltransferase, and Diacylglycerol Acyltransferase on lipid accumulation and composition in Neochloris oleoabundans
'Frontiers Media SA', 2019Co-Authors: Muñoz, Camilo F., D'adamo Sarah, Weusthuis, Ruud A., Wijffels, Rene HubertusAbstract:Microalgal lipids are promising feedstocks for food and biofuels. Since lipid production by microalgae is not yet economically feasible, genetic engineering is becoming a promising strategy to achieve higher lipid accumulation and productivities. Enzymes involved in the Kennedy pathway such as glycerol-3-phosphate acyltransferase (GPAT), lysophosphatidic acid acyltransferase (LPAT), and diacylglycerol acyltransferase (DGAT) catalyze key steps in the formation of triacylglycerol, which is the main constituent of lipids in N. oleoabundans. The overexpression of these enzymes in the targeted strain has a great potential to further increase their triacylglycerol content. We overexpressed single and multiple encoding genes for LPAT, GPAT, and DGAT from Acutodesmus Obliquus in N. oleoabundans. Strains overexpressing single genes produced up to 52% and 45% g · gDW-1, which corresponds to 1.3- and 1.4-fold increase in total fatty acids and triacylglycerols, respectively. The orchestrated expression of the three genes resulted in 49% and 39% g · gDW-1, which is 1.2-folds increase in total fatty acids and triacylglycerols. Single expression of LPAT, GPAT, and DGAT genes resulted in higher lipid productivities during starvation without a significant effect on growth and photosynthetic activity during replete conditions. On the other hand, the simultaneous expression of LPAT, GPAT, and DGAT genes resulted in 52% lower growth rate, 14% lower photosynthetic activity and 4-folds increase in cell diameter. Moreover, the multigene expressing line showed a decrease in carbohydrates and protein content and an increase in pigments during nitrogen starved condition. The single and multiple expression of heterologous genes LPAT, GPAT, and DGAT showed to significantly enhanced the lipid accumulation in N. oleoabundans. Single gene expression resulted in higher lipid production and productivities without having a significant impact in the physiological status of the strains. This approach shows the potential for the generation of microalgal strains with higher economical potential for the production of lipids.publishedVersio
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Improved DNA/protein delivery in microalgae – A simple and reliable method for the prediction of optimal electroporation settings
2018Co-Authors: Muñoz, Camilo F., Sturme, Mark H.j., Martens, Dirk E., Lenny De Jaeger, Lip, Ka Y.f., Olijslager, Justus W.j., Springer Jan, Wolbert, Emil J.h., Eggink Gerrit, Weusthuis, Ruud A.Abstract:Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus Obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.
Mark H. J. Sturme - One of the best experts on this subject based on the ideXlab platform.
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Stable transformation of the green algae Acutodesmus Obliquus and Neochloris oleoabundans based on E. coli conjugation
Algal Research-Biomass Biofuels and Bioproducts, 2019Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Sarah D'adamo, Ruud A WeusthuisAbstract:Abstract Microalgae are an ideal platform for the production of high-value chemicals, nutritional products and biofuels. Genetic engineering could speed up the development of microalgae derived products and reduce the overall production costs. Genetic methods such as particle bombardment, electroporation, Agrobacterium tumefaciens mediated transformation (ATMT), and agitation with glass beads and silicon carbide whiskers have been developed for the genetic transformation of microalgae. However, the transformation efficiency is species dependent, so a variety of transformation methods are required to engineer a wide range of microalgae species. The oleaginous microalgae Acutodesmus Obliquus and Neochloris oleoabundans have a great potential as production platforms due to their ability to produce large amounts of triacylglycerol (TAG). Genetic modification techniques however are required to increase TAG levels further or to modify the fatty acid composition. Recently, a conjugation-based method for the delivery of episomes from bacteria to diatom microalgae has been reported. In this study, we have achieved the successful transformation of green oleaginous microalgal strains by transferring an expression vector via conjugation from E. coli. Since delivery of exogenous DNA into the microalgae cells is only the first step in obtaining transgenic microalgae, we further analyzed transformation efficiencies by PCR and expression of the Clover fluorescent protein in the targeted species.
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improved dna protein delivery in microalgae a simple and reliable method for the prediction of optimal electroporation settings
Algal Research-Biomass Biofuels and Bioproducts, 2018Co-Authors: Camilo F. Muñoz, Mark H. J. Sturme, Lenny De Jaeger, Ka Y F Lip, Justus W J Olijslager, Jan Springer, Emil J H Wolbert, Dirk E Martens, Gerrit Eggink, Ruud A WeusthuisAbstract:Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus Obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.