We attempted to establish a quantitative method to quantify the presence of cyanobacteria in environmental samples. As DNA isolations are performed with commercial kits, it was necessary to evaluate the isolation efficiency of each kit. Therefore, pure cultures and environmental samples were used. The pure cultures were grown in phosphate-free media, where it was meant to consume their own genetic material as a source of phosphate. The latter would result in monoploidy in which the isolation efficiency could be calculated based on cell counts and isolated DNA. For the environmental samples, we used an artificially designed DNA sequence not found in our samples, which was added either before or after DNA isolation, and checked the efficiency of DNA isolation by comparing the qPCR results. The results showed we have higher qPCR inhibition in concentrated samples of pure cultures. With the commercial kit for biofilm isolation, we obtained sufficient results, with isolation efficiencies ranging between 17 and 43%. To determine the isolation efficiency with a commercial kit for planktonic samples, the experiments would need to be repeated as we obtained a remarkable scatter of results. The DNA isolation efficiency from pure culture Synechococcus was approximately 46%, from pure culture Anabaena 0,2% and from planktonic environmental samples up to 2,1%.
|
