title: Agrobacterium tumefaciens mediated transformation of marine microalgae Schizochytrium.
abstract: Schizochytrium was a known docosahexaenoic acid producing marine microalgae. In this study, we have developed a novel transformation approach of Schizochytrium using the Agrobacterium tumefaciens (A. tumefaciens) binary vector system. After co-cultivation of Schizochytrium protoplasts with A. tumefaciens harboring pCAMBIA2301 containing the neomycin phosphotransferase II (NPT II) gene as the selectable marker which confers resistance to G418, the Schizochytrium transformants were successfully obtained on the G418-containing plates. The integration and expression of the transgenes were confirmed by PCR analysis and GUS activity assay. To further validate the transformation system, pCAMBIA2301-EGFP containing the egfp gene was introduced into Schizochytrium. The following results demonstrated that the exogenous egfp gene has been successfully incorporated into the genome of Schizochytrium. In addition, the introduced egfp gene expressed efficiently according to the Western blot and fluorescence assay results. More importantly, the majority of the transformants displayed similar biomass and fatty acid production comparing with the wild type strain. Our results demonstrated that exogenous genes could be expressed efficiently in transgenic Schizochytrium, suggesting that genetically engineered Schizochytrium could be explored by this system.
authors.affiliation: Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, State Oceanic Administration, 178 Daxue Road, Xiamen 361005, Fujian Province, People's Republic of China.
authors.forename: Rubin
authors.initials: R
authors.lastname: Cheng
authors.forename: Ruijuan
authors.initials: R
authors.lastname: Ma
authors.forename: Ke
authors.initials: K
authors.lastname: Li
authors.forename: Hui
authors.initials: H
authors.lastname: Rong
authors.forename: Xiangzhi
authors.initials: X
authors.lastname: Lin
authors.forename: Zhaokai
authors.initials: Z
authors.lastname: Wang
authors.forename: Shanjun
authors.initials: S
authors.lastname: Yang
authors.forename: Yong
authors.initials: Y
authors.lastname: Ma
mesh_terms: D016960:Agrobacterium tumefaciens / Q000737:chemistry / Q000235:genetics*; D018533:Biomass; D015153:Blotting, Western; D004269:DNA, Bacterial / Q000235:genetics*; D005227:Fatty Acids / Q000032:analysis / Q000096:biosynthesis; D017930:Genes, Reporter; D005818:Genetic Engineering; D005822:Genetic Vectors / Q000737:chemistry*; D005966:Glucuronidase; D019868:Kanamycin Kinase / Q000235:genetics* / Q000378:metabolism; D058086:Microalgae / Q000235:genetics* / Q000378:metabolism; D030781:Organisms, Genetically Modified / Q000235:genetics; D011523:Protoplasts / Q000378:metabolism*; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D004269:DNA, Bacterial; D005227:Fatty Acids; D019868:Kanamycin Kinase; D005966:Glucuronidase