title: Development of a nuclear transformation system for Oleaginous Green Alga Lobosphaera (Parietochloris) incisa and genetic complementation of a mutant strain, deficient in arachidonic acid biosynthesis.
abstract: Microalgae are considered a promising source for various high value products, such as carotenoids, ω-3 and ω-6 polyunsaturated fatty acids (PUFA). The unicellular green alga Lobosphaera (Parietochloris) incisa is an outstanding candidate for the efficient phototrophic production of arachidonic acid (AA), an essential ω-6 PUFA for infant brain development and a widely used ingredient in the baby formula industry. Although phototrophic production of such algal products has not yet been established, estimated costs are considered to be 2-5 times higher than competing heterotrophic production costs. This alga accumulates unprecedented amounts of AA within triacylglycerols and the molecular pathway of AA biosynthesis in L. incisa has been previously elucidated. Thus, progress in transformation and metabolic engineering of this high value alga could be exploited for increasing the efficient production of AA at competitive prices. We describe here the first successful transformation of L. incisa using the ble gene as a selection marker, under the control of the endogenous RBCS promoter. Furthermore, we have succeeded in the functional complementation of the L. incisa mutant strain P127, containing a mutated, inactive version of the delta-5 (Δ5) fatty acid desaturase gene. A copy of the functional Δ5 desaturase gene, linked to the ble selection marker, was transformed into the P127 mutant. The resulting transformants selected for zeocine resistant, had AA biosynthesis partially restored, indicating the functional complementation of the mutant strain with the wild-type gene. The results of this study present a platform for the successful genetic engineering of L. incisa and its long-chain PUFA metabolism.
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Boris
authors.initials: B
authors.lastname: Zorin
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Omer
authors.initials: O
authors.lastname: Grundman
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Inna
authors.initials: I
authors.lastname: Khozin-Goldberg
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Stefan
authors.initials: S
authors.lastname: Leu
authors.affiliation: Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel.
authors.forename: Michal
authors.initials: M
authors.lastname: Shapira
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Yuval
authors.initials: Y
authors.lastname: Kaye
authors.affiliation: UMR 7141 CNRS/Université Pierre et Marie Curie, Institut de Biologie Physico-Chimique, Paris, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Tourasse
authors.affiliation: UMR 7141 CNRS/Université Pierre et Marie Curie, Institut de Biologie Physico-Chimique, Paris, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Vallon
authors.affiliation: Microalgal Biotechnology Laboratory, French Associates Institute for Agriculture and Biotechnology of Drylands, J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion, Israel.
authors.forename: Sammy
authors.initials: S
authors.lastname: Boussiba
mesh_terms: D016718:Arachidonic Acid / Q000172:deficiency / Q000378:metabolism*; D000460:Chlorophyta / Q000235:genetics / Q000378:metabolism*; D015525:Fatty Acids, Omega-3 / Q000378:metabolism; D043371:Fatty Acids, Omega-6 / Q000378:metabolism
chemical_list: D015525:Fatty Acids, Omega-3; D043371:Fatty Acids, Omega-6; D016718:Arachidonic Acid