Heterotrophic Bacterial Production

Methods

Heterotrophic bacterial production method (2007-2008)

For each sample, 1.7 mL seawater were incubated with approximately 20 nM 3H-Leucine for one hour. Samples were done in triplicate with duplicate TCA-killed controls. After the incubation was complete, all samples were killed with an addition of 100% TCA for a final concentration of 5% TCA. The samplers were filtered on 0.2 µm pore size polycarbonate filters and three additions of 1 mL 5% TCA were made. The filters were dried in individual scintillation vials at room temperature before scintillation cocktail was added and analyzed. Samples were analyzed for disintegrations per minute on a Beckan LS6000A liquid scintillation counter. Disintegrations per minute were converted to protein synthesis rates assuming 3.1 kg C mol-1 leucine and 24 h day-1 (Simon & Azam, 1989). This calculation can be normalized by the corresponding cell abundance measurement when possible.

Heterotrophic bacterial production method (2014-current)

For each sample, 1.7 mL seawater were incubated with approximately 20 nM 3H-Leucine for one hour. Samples were done in triplicate with single TCA-killed controls. After the incubation was complete, all samples were killed with an addition of 100% TCA for a final concentration of 5% TCA. The samples were processed using the centrifugation method as described in Smith et al. (1992). Samples were frozen and stored at -20°C as needed. Samples were analyzed for disintegrations per minute on a Beckan LS6000A liquid scintillation counter. Disintegrations per minute were converted to protein synthesis rates assuming 3.1 kg C mol-1 leucine and 24 h day-1 (Simon & Azam, 1989). This calculation can be normalized by the corresponding cell abundance measurement when possible.