Samples for the measurement of nitrate plus nitrite and phosphate (PO43−) concentrations were filtered through precombusted GF/F filters and analyzed using standard techniques on a Lachat QuikChem 8000 Flow Injection Analyzer. Chl a samples from each depth were filtered onto GF/F filters, extracted in the dark at 3 °C in 90% acetone for 24 h and measured fluorometrically using a Turner Fluorometer TD-700 as described in Welschmeyer et al.

For DNA collection and extraction, seawater was filtered through SterivexTM filters using gentle peristaltic pumping and flash-frozen in liquid N2. DNA was extracted using the DNeasy Plant Kit (Qiagen, Germantown, MD) using modifications to the manufacturer’s guidelines described in detail in Moisander et al. 2007. On-column steps were automated using a QIACube (Qiagen). DNA was quantified using the Picogreen® dsDNA Quantitation kit (Molecular Probes, Eugene, OR).

Gene-based abundance estimates of UCYN-A1, UCYN-A2, Crocosphaera (UCYN-B), Trichodesmium, Richelia associated with Hemiaulus (Het-2), and gamma A (γ-24774A11) were determined using Taqman® qPCR assays. Protocols used for all aspects of qPCR analysis, including reaction conditions, the use of linearized plasmids and inhibition reactions, and calculation of unknowns follow those described in detail by Goebel et al. 2010, apart from a 64 °C annealing temperature for the UCYN-A2 assay. The LOD and LOQ for all assays ranged between 25-31 and 200-250 nifH copies l−1, respectively. Targets with nifH copies >LOD and <LOQ are detected not quantified (DNQ).

Methodology described in depth in Turk-Kubo et al. (2021).