Molecular probe sets for the detection of toxic algae for use in sandwich hybridization formats

Abstract

Molecular probes can be used for early and rapid detection of toxic algal species. The sandwich hybridization requires two probes for each species, a capture probe and a nearly adjacent signal probe. Probe sets for the species-specific identification of the toxic algal species Gymnodinium catenatum, Protoceratium reticulatum, Lingulodinium polyedrum, Prymnesium parvum, Chrysochromulina polylepis, Pseudo-nitzschia multiseries, Pseudo-nitzschia australis, Pseudo-nitzschia seriata and Pseudo-nitzschia pungens were designed. A genus probe set for Pseudo-nitzschia species was adapted, and all probe sets were tested for specificity. The target molecules for the probe sets are the large and the small subunit ribosomal RNAs (rRNAs). The specificity of the different probe sets was tested using a sandwich hybridization assay in a microtiter plate with rRNA isolated from laboratory strains of the target species and closely related species. The assay showed that eight probe sets were highly specific. For G. catenatum and P. parvum, the probe sets detected one non-target species outside the target species. These 10 probe sets are valuable tools for identifying and monitoring different toxic algae. The microtiter plate assay is a cheap and effective means for testing probe specificity.