Culturing and experimental conditions
Experimental cultures were grown with a semi-continuous culturing method at 28 degrees C in autoclave-sterilized artificial seawater medium with nutrients added in concentrations equivalent to the recipe for the Aquil medium (except for NO3-), as in Garcia et al. (2011) and originally described by Morel et al. (1979).

Light experiment and cellular growth rates
Triplicate cultures were grown in 800 mL polystyrene flasks under 5 irradiances (18, 40, 100, 180, 300 umol quanta per m^2 per second) and diluted every 2-3 days to 10-20 x 103 cells per mL. Cells were counted microscopically in each replicate culture with a hemocytometer at the end of each dilution period, and steady state growth rates were calculated from an increase in culture cell number per unit volume between 2-3 dilution periods (4-6 days) after cultures were acclimated to treatment conditions for 7-10 generations. To calculate growth rates, the investigators used the equation N_T=N₀e^µT, where N₀ and N_T are the initial and final culture cell densities, respectively and T is the amount of time in days between culture cell number estimates. With this method, the dilution rate is determined by the growth rate of the algae as determined by the experimental treatments, rather than by controlling the growth rate through imposing a dilution rate, as one does for continuous cultures.

Cell diameters of ~12 cells from treatment replicates were measured with an ocular micrometer. In the light experiment, cells in one replicate from each light were measured treatment twice, once in the middle of the light period and once at the end of the light period on the same day.

Light was supplied on a 12:12 light:dark cycle with cool white fluorescent bulbs. The investigators terminally sampled each replicate culture 24 hours after the last dilution for N2-fixation rates and CO2-fixation rates.

References:
Garcia, N. S., F.-X. Fu, , C. L. Breene, P. W. Bernhardt, M. R. Mulholland, J. A. Sohm, and D. A. Hutchins. 2011. Interactive effects of irradiance and CO2 on CO2- and N2 fixation in the diazotroph Trichodesmium erythraeum (Cyanobacteria). J. Phycol. 47: 1292-1303. DOI: 10.1111/j.1529-8817.2011.01078.x

Morel, F. M. M., J. G. Rueter, D. M. Anderson, and Guillard, R. R. L. 1979. Aquil: Chemically defined phytoplankton culture medium for trace metal studies. J. Phycol. 15:135-141. DOI: 10.1111/j.1529-8817.1979.tb02976.x