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EU μAQUA made early warning systems for freshwater pathogens and toxins. Barcodes for each cyanobacterial toxin gene published before 2011 were designed and used in a microarray format to capture messenger RNA to detect toxin gene expression at early stages of bloom development. A reverse transcriptase (RT) microarray was developed to detect toxin expression, which had low expression levels. Probes immobilized on the microarray slide captured the mRNA and were extended directly on the microarray. RT extension incorporated fluorescently labeled oligonucleotides to ensure a high signal detected by the microarray scanner. The CYANO RT microarray was laboratory tested with known toxic cyanobacteria and field-tested. Hybridizations without RT extensions were barely detectable in the cultured strains. However, with RT extensions, hybridized mRNA was easily detected. Field samples were equally successful and consistent with companion studies from the same sites using HPLC/(MS-MS) (High Performance Liquid Chromotography/ Mass Spec). In some cases, amplified expression produced a signal with no detection of that toxin using chemical means. The RT microarray may be more sensitive than HPLC/MS-MS. Further studies are needed to determine if the RT-microarray is detecting a very low expression of the toxin genes and, hence, more sensitive as an early warning system predicting the toxin potential.
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