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Engineering a marine microalga Chlorella sp. as the cell factory.

ABSTRACT: The use of marine microalgae in industrial systems is attractive for converting CO₂ into value-added products using saline water and sunlight. The plant nature and demonstrated industrial potential facilitate Chlorella spp. as excellent model organisms for both basic research and commercial application. However, the transformation method has not been developed in marine Chlorella spp., thus genetic engineering is hindered in exploiting the industrial potentialities of these strains. In this study, we provided a transformation protocol for the marine Chlorella strain MEM25, which showed robust characteristics, including high production of proteins and polyunsaturated fatty acids in multiple cultivation systems over various spatial-temporal scales. We showed that transformants could be obtained in a dramatically time-saving manner (comparable to Saccharomyces cerevisiae) with four functional proteins expressed properly. The transgenes are integrated into the genome and can be successfully inherited for more than two years. The development of a marine Chlorella transformation method, in combination with the complete genome, will greatly facilitate more comprehensive mechanism studies and provide possibilities to use this species as chassis for synthetic biology to produce value-added compounds with mutual advantage in neutralization of CO₂ in commercial scales.

SUBMITTER: Gu X

PROVIDER: S-EPMC10485975 | biostudies-literature | 2023 Sep

REPOSITORIES: biostudies-literature

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Engineering a marine microalga Chlorella sp. as the cell factory.

Gu Xinping X Deng Ying Y Wang Aoqi A Gan Qinhua Q Xin Yi Y Paithoonrangsarid Kalyanee K Lu Yandu Y

Biotechnology for biofuels and bioproducts 20230907 1

The use of marine microalgae in industrial systems is attractive for converting CO<sub>2</sub> into value-added products using saline water and sunlight. The plant nature and demonstrated industrial potential facilitate Chlorella spp. as excellent model organisms for both basic research and commercial application. However, the transformation method has not been developed in marine Chlorella spp., thus genetic engineering is hindered in exploiting the industrial potentialities of these strains. I ...[more]

PMID: 37679828

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Project description:Over the last decades, microalgae have gained popularity due to demand for novel environmental green solutions and development of innovative mass-production sources for multiple processes, including animal feed and human diet, turning microalgae into an exquisite candidate for several ecofriendly technologies. Notwithstanding, there is a catch. Most species of microalgae, as the case of common Chlorella vulgaris (C. vulgaris) display a recalcitrant cell wall, characterized by a complex matrix of polysaccharides and glycoproteins, which constitutes a major barrier for monogastric species digestibility and extraction of inner valuable nutritional compounds. To overcome this limitation, the development of feed enzymes, in particular Carbohydrate-Active enZymes (CAZymes) with capacity to disrupt C. vulgaris cell wall may contribute to improve the bioavailability of these microalgae compounds in monogastric diets, namely at high levels of incorporation. In order to disclosure novel combination of feed enzymes to disrupt C. vulgaris cell wall, a lab protocol was implemented by our research team containing the following key steps: after microalgae cultivation and having available a repertoire of two hundred pre-selected CAZymes produced by high-throughput technology, the step 1 is the individual screening of the most functional enzymes on disrupting C. vulgaris cell wall (versus a control, defined as the microalgae suspension incubated with PBS) and the determination of reducing sugars released by the 3,5-dinitrosalicylic acid (DNSA) method; step 2 concerns on finding the best CAZymes cocktail, testing the synergistic effect of enzymes, to disrupt C. vulgaris cell wall (in parallel with running the control) along with characterization of each enzyme thermostability and resistance to proteolytic attack, to which feed enzymes are subjected in the animal gastrointestinal tract; step 3 is the assessment of C. vulgaris cell wall degradation degree by measuring the amount of reducing sugars released by the DNSA method, fatty acid analysis by gas chromatography (GC) with flame ionization detector (FID), oligosaccharides quantification by high performance liquid chromatography (HPLC) equipped with an electrochemical detector (ECD), protein content by the Kjeldahl method, and various pigments (chlorophylls a and b, and total carotenoids) in the supernatant. In the correspondent residue, we also assessed cellular counting using a Neubauer chamber by direct observation on a bright-field microscope and fluorescence intensity, after staining with Calcofluor White for both control and CAZymes cocktail treatments, on a fluorescence microscope. Beyond animal feed industry with impact on human nutrition, our lab protocol may increase the yield in obtaining valued constituents from C. vulgaris microalga for other biotechnological industries.

Dataset Information

Engineering a marine microalga Chlorella sp. as the cell factory.

Publications

Engineering a marine microalga Chlorella sp. as the cell factory.

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