As an important component of dissolved organic matter (DOM), nucleic acids (DNA and RNA) are essential nutrient and energy sources in aquatic microbial food web. Therefore, it is important to understand the bioavailability of nucleic acids. The bioavailability of nucleic acids was investigated by a batch of incubation experiments, adding fish DNA and yeast RNA into water samples with different salinity collected from the Yangtze River estuary in the spring of 2012. According to the results, 20%-50% of dissolved DNA was transformed into particulate DNA quickly with the conversion rates increasing with salinity, only 10% dissolved RNA was transformed into particulate RNA and the salinity had no effect on the conversation rates. In each incubation experiment, the microbial utilization kinetic curves of dissolved nucleic acids were fitted to the Sigmoid model. There were lag periods of 30-80 hours followed by the rapid utilization phase and then the stagnation phase. The results also showed that the bacteria in seawater had higher maximum utilization rate than the bacteria in estuarine and fresh water. Dissolved nucleic acids spiked in estuarine water can be bound to colloids and particles at some extent, only those free dissolved or enzymatically hydrolysable forms are bioavailable. The percentage of bioavailable RNA (80%-90%) was significantly higher than that of bioavailable DNA and it did not change significantly with salinity while the percent of bioavailable DNA decreased from 78% to 50% with salinity. Therefore, the speciation and bioavailability are significantly different between DNA and RNA across the estuarine salinity gradient.