Marine nitrogen (N2) fixation was historically considered to be absent or reduced in nitrate (NO3−) rich environments. This is commonly attributed to the lower energetic cost of NO3− uptake compared to diazotrophy in oxic environments. This paradigm often contributes to making inferences about diazotroph distribution and activity in the ocean, and is also often used in biogeochemical ocean models. To assess the general validity of this paradigm beyond the traditionally used model organism Trichodesmium spp., we grew cultures of the unicellular cyanobacterium Crocosphaera watsonii WH8501 long term in medium containing replete concentrations of NO3−. NO3− uptake was measured in comparison to N2 fixation to assess the cultures’ nitrogen source preferences. We further measured culture growth rate, cell stoichiometry, and carbon fixation rate to determine if the presence of NO3− had any effect on cell metabolism. We found that uptake of NO3− by this strain of Crocosphaera was minimal in comparison to other N sources (~2–4% of total uptake). Furthermore, availability of NO3− did not statistically alter N2 fixation rate nor any aspect of cell physiology or metabolism measured (cellular growth rate, cell stoichiometry, cell size, nitrogen fixation rate, nitrogenase activity) in comparison to a NO3− free control culture. These results demonstrate the capability of a marine diazotroph to fix nitrogen and grow independently of NO3−. This lack of sensitivity of diazotrophy to NO3− suggests that assumptions often made about, and model formulations of, N2 fixation should be reconsidered.