DETERMINATION OF RNA AND DNA CONCENTRATIONS IN NATURAL PLANKTON SAMPLES USING THIAZOLE ORANGE IN COMBINATION WITH DNASE AND RNASE DIGESTIONS

A fluorometric technique, based on the combination of RNase and DNase incubation with the use of thiazole orange (RNase / DNase method), was investigated to determine DNA and RNA concentrations in marine plankt on. Tests were performed to optimize both RNase and DNase assay condit ions. The RNase assay should be conducted at 37 degrees C for 20 min w ith 0.5 mu g . mL(-1) of DNase-free RNase. An incubation at 25 degrees C for 20 min with 10 units . mL(-1) of RNase-free DNase were the opti mal conditions required for DNA digestion by DNase. The detection limi ts in terms of minimum biomass for reliable measurements of DNA and RN A were 7.5 and 10 mu g of protein .(mL assay)(-1), respectively. RNA a nd DNA concentrations were estimated in oligotrophic water samples usi ng the RNase / DNase and other available methods (e.g. a double fluoro chrome method). The different techniques provided similar DNA estimati ons. However, the RNase / DNase method provided the highest sensitivit y and a low variability for the estimation of RNA.