Outbreaks of toxic cyanobacteria have been increasing on the Great Lakes. Cyanobacteria in these toxic blooms can produce a family of hepatotoxic peptides, called microcystins. There are two hypotheses surrounding the origin of microcystin production in Lake Ontario: (1) it originates offshore and is circulated throughout the lake via water currents and (2) it is created in eutrophic embayments and is transported to the main lake. To investigate the existence and origin of microcystin production in Lake Ontario, samples were collected in embayments along the southern and eastern shores and from open water. Molecular analysis by PCR revealed the potential for microcystin production, which was then compared to actual microcystin production determined via the biochemical protein phosphatase inhibition assay (PPIA). This information was then used to evaluate the relative contribution of embayments vs. offshore regions to microcystin production in Lake Ontario.