Microalgae serve as a promising source for the production of biofuels and bio-based chemicals. They are superior to terrestrial plants as feedstock in many aspects and their biomass is naturally rich in lipids, carbohydrates, proteins, pigments, and other valuable compounds. Due to the relatively slow growth rate and high cultivation cost of microalgae, to screen efficient and robust microalgal strains as well as genetic modifications of the available strains for further improvement are of urgent demand in the development of microalgae-based biorefinery. In genetic engineering of microalgae, transformation and selection methods are the key steps to accomplish the target gene modification. However, determination of the preferable type and dosage of antibiotics used for transformant selection is usually time-consuming and microalgal-strain-dependent. Therefore, more powerful and efficient techniques should be developed to meet this need. In this review, the conventional and emerging genome-editing tools (e.g., CRISPR-Cas9, TALEN, and ZFN) used in editing the genomes of nuclear, mitochondria, and chloroplast of microalgae are thoroughly surveyed. Although all the techniques mentioned above demonstrate their abilities to perform gene editing and desired phenotype screening, there still need to overcome higher production cost and lower biomass productivity, to achieve efficient production of the desired products in microalgal biorefineries.
β-carotene is a valuable pigment abundant in some microalgal species but the low β-carotene productivity of microalgae has become the major obstacles against its commercialization. This work aims to improve the productivity of algae-based β-carotene via genetic engineering approaches. First, a synthetic psy gene construct (891 bp) encoding 297 amino acids is expressed in Scenedesmus sp. CPC2 host to enhance the β-carotene production. The synthetic psy gene is designed by considering the highest consensus of amino acids (i.e., 62% identity) from Chlamydomonas reinhardtii, Dunaliella salina, and Mariella zofingiensis. The original β-carotene content in wild-type Scenedesmus sp. CPC2 is 10.8 mg g -cell when grown on BG11 medium under 2% CO aeration, 150 μmol m s light intensity and 25°C. After transformation of the psy gene into the microalgal host, the β-carotene content of the best recombinant strain (i.e., transformant CPC2-4) significantly increased to over 30 mg g -cell. The optimal production of β-carotene with the CPC2-4 recombinant strain was achieved when the strain is grown on BG11 medium amended with 0.075 g of MgSO , giving approximately 3-fold higher β-carotene content than that of the wild-type strain. The best cellular β-carotene content obtained (i.e., 31.8 mg g ) is superior to most algae-based β-carotene production performance reported in the literature.
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