The zooplankton assemblage was collected using a 1m2 Multiple Opening/Closing Nets and Environmental Sampling System (MOCNESS), with all nets and cod ends at 202 µm mesh. All tows were conducted at Station ALOHA (22° 45’N 158° W) A total of 5 tows sampled the full water column between 1000 m and the sea surface, with 3 day and 2 night paired tows. Depth strata were as follows: 0-1000m (Net 0), 1000-800 m (Net 1), 800-600m (Net 2), 600-500m (Net 3), 500-400m (Net 4), 400-300m (Net 5), 300-200m (Net 6), 200-100m (Net 7), 100-50m (Net 8), 50-0m (Net 9).
Bulk plankton from each net and tow were quantitatively split using a Folsom plankton splitter, with additional processing or preservation for several analyses. Two additional ‘short’ MOCNESS tows were conducted to sample the base of the mesopelagic (700-1000m) and epipelagic (0-200 m) for metatranscriptomic analyses of the zooplankton assemblage. Rapid recovery and preservation of the material was important in this case, given primary interest in gene expression profiles, and so the nets were flown as quickly as possible through midwater.
Quantitative subsamples of bulk plankton were preserved as follows:
- Formalin-preserved – sodium tetraborate buffered formaldehyde, preserved broadly following CalCOFI protocols (https://calcofi.org/about-calcofi/calcofi-handbook/301-net-sample-preservation.html). Analyses: ZooScan, Microscopy
- Size-fractionated into 5 or 2 fractions (0.2-0.5 mm, 0.5-1.0 mm, 1.0-2.0 mm, 2.0-5.0 mm, >5.0 mm OR 0.2-1.0 mm, >1.0 mm), filtered onto 200 µm Nitex filters, rinsed with ammonium formate, and followed by cryopreservation and storage at -80° C. Analyses: biomass, bioelements, isotopes.
- Filtered onto 200 µm Nitex filters, followed by preservation in RNALater and storage at -80° C. Analyses: Metabarcoding.
- Filtered onto 200 µm Nitex filters, flash frozen and stored in liquid nitrogen. Analyses: Electron Transport System (ETS) activity assay.
- Size-fractionated into 2 fractions (0.2 - 1.0 mm, > 1.0 mm) and analyzed shipboard for alkaline phosphatase activity (APA assay)