Development of a stable transformation method for Saprolegnia parasitica

Abstract

Saprolegniosis in salmonids, a disease caused by the oomycete Saprolegnia parasitica, poses a serious global threat to wild salmon and to aquaculture. To be able to functionally characterise genes in S. parasitica, it is essential to develop a stable transformation method for S. parasitica. We describe for the first time a method that can generate stable transgenic S. parasitica strains. Transformants were generated following the uptake and integration of a mutated gene from Achlya hypogyna conferring imidazole resistance, CYP51 using S. parasitica protoplasts in the presence of polyethylene glycol (PEG) and lipofectamine. This leads to production of CYP51 protein which catalyses a crucial demethylation step in the biosynthesis of ergosterol. As a result, there is no disruption of ergosterol synthesis and the transformants, but not the wild type S. parasitica, can grow in the presence of imidazole. Putative transformants growing in the presence of up to 10 mM imidazole were confirmed by PCR.