GENETIC-TRANSFORMATION OF DINOFLAGELLATES (AMPHIDINIUM AND SYMBIODINIUM) - EXPRESSION OF GUS IN MICROALGAE USING HETEROLOGOUS PROMOTER CONSTRUCTS

Genetic transformation of two dinoflagellates (Amphidinium sp., Symbio dinium microadriaticum) was achieved using plasmid constructs containi ng the neomycin phosphotransferase gene (nptll) fused to the Agrobacte rium nos promoter, or the hygromycin B phosphotransferase gene (hpt) f used to the bidirectional Agrobacterium p1'2' promoter. Gene transfer into intact (walled) dinoflagellate cells was achieved by agitation in the presence of silicon carbide (SiCa) whiskers. Transformation rates of 5-24 transformants per 10(7) cells were obtained. Southern hybridi zation of transformants revealed stable integration of multiple copies of the constructs. Activity of integrated copies of the beta-glucoron idase (GUS) reporter gene coupled to the cauliflower mosaic virus 35S promotor or the p1'2' promoter was confirmed both histochemically and fluorometrically. This is the first report of successful application b f heterologous and widely used promoter and reporter genes in microalg ae, and is the first demonstration of transformation of a dinoflagella te. There appear to be no substantial barriers to transformation of Am phidinium and Symbiodinium, which must now be considered as the first of the dinoflagellate genera accessible to genetic manipulation.