The genetic intractability of Symbiodinium microadriaticum to standard algal transformation methods

• Published in: PloS one 2019-02, Vol.14 (2), p.e0211936-e0211936
• Main Authors: Chen, Jit Ern, Barbrook, Adrian C, Cui, Guoxin, Howe, Christopher J, Aranda, Manuel
• Format: Article
• Language: English
• Subjects: Algae; Animals; Anthozoa - microbiology; Antibacterial agents; Antibiotics; Atrazine; Atrazine - pharmacology; Biochemistry; Biology and Life Sciences; Carbides; Cell Nucleus - genetics; Cell Nucleus - metabolism; Chloramphenicol; Chloramphenicol - pharmacology; Chloromycetin; Chloroplast Proteins - genetics; Chloroplast Proteins - metabolism; Chloroplasts; Chloroplasts - genetics; Chloroplasts - metabolism; Coral reefs; Corals; CRISPR; Deoxyribonucleic acid; Dinoflagellates; Dinoflagellida - genetics; Dinoflagellida - metabolism; DNA; DNA methylation; Drug Resistance - genetics; Editing; Electroporation; Gene expression; Genes; Genetic aspects; Genetic research; Genetic transformation; Genetic vectors; Genome editing; Genome-wide association studies; Genomes; Genomics; Herbicides; Mathematical analysis; Medicine and Health Sciences; Microorganisms; Neomycin; Nuclear transformations; Proteins; Research and analysis methods; Silicon; Silicon carbide; Silicon carbides; Silicon compounds; Stem cells; Symbiodinium; Symbiodinium microadriaticum; Symbiosis; Transformation, Genetic
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0211936

Modern transformation and genome editing techniques have shown great success across a broad variety of organisms. However, no study of successfully applied genome editing has been reported in a dinoflagellate despite the first genetic transformation of Symbiodinium being published about 20 years ago. Using an array of different available transformation techniques, we attempted to transform Symbiodinium microadriaticum (CCMP2467), a dinoflagellate symbiont of reef-building corals, with the view to performing subsequent CRISPR-Cas9 mediated genome editing. Plasmid vectors designed for nuclear transformation containing the chloramphenicol resistance gene under the control of the CaMV p35S promoter as well as several putative endogenous promoters were used to test a variety of transformation techniques including biolistics, electroporation and agitation with silicon carbide whiskers. Chloroplast-targeted transformation was attempted using an engineered Symbiodinium chloroplast minicircle encoding a modified PsbA protein expected to confer atrazine resistance. We report that we have been unable to confer chloramphenicol or atrazine resistance on Symbiodinium microadriaticum strain CCMP2467.