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Projeto de investigação
Development of novel microalgal mutants with improved pigmentation and protein contents for industrial waste-based fermentation
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Improving the heterotrophic media of three Chlorella vulgaris mutants toward optimal color, biomass and protein productivity
Publication . Trovão, Mafalda; Cunha, Miguel; Santo, Gonçalo Espírito; Pedroso, Humberto; Reis, Ana; Barros, Ana; Correia, Nádia; Schüler, Lisa; Costa, Monya; Ferreira, Sara; Cardoso, Helena; Ventura, Márcia; Varela, João; Silva, Joana; Freitas, Filomena; Pereira, Hugo; DQ - Departamento de Química; UCIBIO - Applied Molecular Biosciences Unit; LAQV@REQUIMTE; Nature Publishing Group
The high production costs and unappealing sensory properties still limit the widespread commercialization of microalgae feedstocks. Therefore, this work focused on fine-tuning the heterotrophic medium composition to cultivate novel green, yellow, and white Chlorella vulgaris mutant strains. Screening assays were carried out to select the most significant factors, and different nutrient concentrations were optimized by modelling biomass and protein productivity, specific growth rate, and color, via response surface methodology. The biomass and protein productivities achieved by these strains were improved by up to 70% and 94%, respectively. Additionally, biomass color was correlated with medium composition for the first time, allowing the improvement of the yellow and white mutant colorations by 20%. Overall, the findings of this study are vital to overcoming the challenges of the biobased industry, allowing the enhancement of the cost-effectiveness, attractiveness, and nutritional profiles of microalgae-based products in different markets and applications.
Isolation and Selection of Protein-Rich Mutants of Chlorella vulgaris by Fluorescence-Activated Cell Sorting with Enhanced Biostimulant Activity to Germinate Garden Cress Seeds
Publication . Trovão, Mafalda; Schüler, Lisa; Pedroso, Humberto; Reis, Ana; Santo, Gonçalo Espírito; Barros, Ana; Correia, Nádia; Ribeiro, Joana; Bombo, Gabriel; Gama, Florinda; Viana, Catarina; Costa, Monya M.; Ferreira, Sara; Cardoso, Helena; Varela, João; Silva, Joana; Freitas, Filomena; Pereira, Hugo; UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química; MDPI - Multidisciplinary Digital Publishing Institute
Microalgae are a promising feedstock with proven biostimulant activity that is enhanced by their biochemical components (e.g., amino acids and phytohormones), which turns them into an appealing feedstock to reduce the use of fertilisers in agriculture and improve crop productivity and resilience. Thus, this work aimed to isolate protein-rich microalgal mutants with increased biostimulant activity. Random mutagenesis was performed with Chlorella vulgaris, and a selection of protein-rich mutants were sorted through fluorescence-activated cell sorting (FACS), resulting in the isolation of 17 protein-rich mutant strains with protein contents 19–34% higher than that of the wildtype (WT). Furthermore, mutant F4 displayed a 38%, 22% and 62% higher biomass productivity, growth rate and chlorophyll content, respectively. This mutant was then scaled up to a 7 L benchtop reactor to produce biomass and evaluate the biostimulant potential of this novel strain towards garden cress seeds. Compared to water (control), the germination index and the relative total growth increased by 7% and 19%, respectively, after the application of 0.1 g L−1 of this bioproduct, which highlights its biostimulant potential.
Oxyfluorfen
Publication . Trovão, Mafalda; Cardoso, Lucas; Schüler, Lisa; Machado, Adriana; Santo, Gonçalo Espírito; Pedroso, Humberto; Reis, Ana; Barros, Ana; Correia, Nádia; Costa, Monya; Ferreira, Sara; Cardoso, Helena; Mateus, Marília; Silva, Joana; Pereira, Hugo; Freitas, Filomena; Varela, João Carlos Serafim; UCIBIO - Applied Molecular Biosciences Unit; Elsevier
Nowadays, there is an increasing demand for novel feedstocks and alternative protein sources to meet global needs. Because of their rich nutritional profiles and high protein contents, microalgae-based food products and supplements are being developed. Nonetheless, these products present organoleptic characteristics such as taste, smell and colour that are often considered unpleasant by human and animal consumers. To address this constraint, strain improvement approaches such as random mutagenesis have been used, which combined with the right selection strategy, lead to more appealing microalgal biomass. In this work, a novel selection strategy using oxyfluorfen, an inhibitor of the chlorophyll synthesis pathway, was applied for the first time to isolate chlorophyll-deficient strains of Scenedesmus rubescens and Chlorella vulgaris upon treatment with ethyl methanesulfonate (EMS). With this approach, one S. rubescens brownish (37Y01) mutant strain, as well as two C. vulgaris mutant strains, one yellow (31Y15) and one white (31W62), were obtained. S. rubescens 37Y01 displayed a reduced protein content of 19.1% dry weight (DW) compared to that of the wildtype, which presented a protein content of 25.0% DW. C. vulgaris wildtype and mutants exhibited higher protein contents, in the 42.8‐44.3% DW range, compared to Scenedesmus rubescens (p < 0.05). The selective pressure of this inhibitor allowed the selection of S. rubescens and C. vulgaris mutants displaying 55% and 95% decrease in chlorophyll content, respectively. The reduced chlorophyll content greatly improves the sensory properties and consumer acceptance of established mutants, increasing the potential of both strains as feedstocks to develop novel food products.
Random Mutagenesis as a Promising Tool for Microalgal Strain Improvement towards Industrial Production
Publication . Trovão, Mafalda; Schüler, Lisa M.; Machado, Adriana; Bombo, Gabriel; Navalho, Sofia; Barros, Ana; Pereira, Hugo; Silva, Joana; Freitas, Filomena; Varela, João; DQ - Departamento de Química; UCIBIO - Applied Molecular Biosciences Unit; MDPI - Multidisciplinary Digital Publishing Institute
Microalgae have become a promising novel and sustainable feedstock for meeting the rising demand for food and feed. However, microalgae‐based products are currently hindered by high production costs. One major reason for this is that commonly cultivated wildtype strains do not possess the robustness and productivity required for successful industrial production. Several strain improvement technologies have been developed towards creating more stress tolerant and productive strains. While classical methods of forward genetics have been extensively used to determine gene function of randomly generated mutants, reverse genetics has been explored to generate specific mutations and target phenotypes. Site‐directed mutagenesis can be accomplished by employing different gene editing tools, which enable the generation of tailor‐made genotypes. Nevertheless, strategies promoting the selection of randomly generated mutants avoid the introduction of foreign genetic material. In this paper, we review different microalgal strain improvement approaches and their applications, with a primary focus on random mutagenesis. Current challenges hampering strain improvement, selection, and commercialization will be discussed. The combination of these approaches with high‐throughput technologies, such as fluorescence‐activated cell sorting, as tools to select the most promising mutants, will also be discussed.
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Entidade financiadora
Fundação para a Ciência e a Tecnologia
Programa de financiamento
OE
Número da atribuição
2020.06814.BD