Dataset: SBDOM11 mini-remineralization expt.
Deployment: PS1103

Dilution batch-culture bioassay experiments were established 11 May 2011 - 15 May 2011 on the SBDOM 11 cruise using water samples collected from the upper 8m of the water column. Water was gravity-filtered from the sampling Niskin bottle through two mixed cellulose ester filters (1.2 µm and 0.2 µm). Duplicate 500 mL polycarbonate bottles were filled with 375 mL 0.2 µm filtrate (filter sterilized media) and 125 mL 1.2 µm filtrate (bacterial inoculum) collected from the same cast. Bottles were incubated in the dark at 11degrees C for 10 weeks. Samples for bacterial abundance, DOM, and community composition by DNA were collected by decanting from the bottle at specified intervals.

DOC was quantified by high-temperature combustion on a modified Shimadzu TOC-V. We used 1.5 µM DOC as a detection limit for these experiments.

Bacterial abundance was measured using a BD LSRII flow cytometer with an autosampler attachment. Samples were preserved in sterile cryovials with 0.2% final concentration paraformaldehyde, then stored frozen until analysis. Samples were stained with SYBR Green I for enumeration.

DNA was collected from duplicate bottles combined (to conserve volume); 125 mL from each bottle was vacuum-filtered onto a 0.2 µm polyethersulfone filter and frozen. Samples were lysed in sucrose lysis buffer with 1% w/v sodium dodecyl sulfate and 0.2 mg mL-1 proteinase-K, and genomic DNA was extracted using a Qiagen DNEasy silica centrifugation kit. DNA was used to conduct multiplex amplicon pyrosequencing using primers 8F and 338R, following the protocol of Nelson et al. (2014; doi: 10.1111/1462-2920.12241). Sequence analysis was conducted in mothur (Schloss et al. 2009; doi: 10.1128/AEM.01541-09) using a non-redundant subset of the SILVA SSU Ref 16S alignment database (v111) curated as in Nelson et al. (2014).