Cold-water coral microbiomes (Lophelia pertusa) from Gulf of Mexico and Atlantic Ocean: raw data

Three biological replicates (individual colonies of L. pertusa) were sampled at four sites: Viosca Knoll 906 (VK906), Viosca Knoll 826 (VK826), West Florida Slope 1 (WFS1), and Atlantic 1 (ATL1). Samples acquired during cruises in August and September 2009 with names prefixed by 3705 or 3731 were collected by the Johnson-Sea-Link submersible (Harbor Branch Oceanographic Institution), using the Kellogg sampler (Kellogg et al., 2009). The sampler’s individual compartments were cleaned at the surface using ethanol, filled with sterile deionized water and sealed. Coral branches were collected, placed into the containers after ambient seawater evacuated the freshwater, and the containers were re-sealed at depth. Samples with names beginning with ROV00 were collected using the remotely-operated vehicle (ROV) Kraken II (University of Connecticut) during a research cruise in September 2010. The ROV carried several individual polyvinylchloride quivers that were cleaned with ethanol, filled with sterile deionized water and sealed at the surface with rubber stoppers. Immediately prior to collection, a quiver was opened, the sample placed inside, and the quiver sealed before the ROV continued its deployment. Upon return to the surface, all L. pertusa samples were transferred to sterile tubes, covered in Thermo Fisher Scientific's RNAlater Stabilization Solution and incubated overnight at 4 degrees Celsius (ºC) to allow the preservative to permeate the coral tissues before transfer to -20º C for long-term storage.

Two polyps from each coral sample (taken from the middle or tip of the branch to avoid any potential contamination at the base where the sampling claw was in contact with the coral) were combined to homogenize the variability of the bacterial community that may exist between polyps. The calyces containing polyps of L. pertusa were broken from the main branch with sterile pliers and placed into sterile aluminum dishes. The calyces were cracked open with a sterile hammer and the tissue was removed from the skeleton using an airbrush with sterile phosphate buffered saline (PBS) and sterile forceps, taking care to minimize dilution of the sample with the PBS. While mainly tissue, the samples may have entrained some coral mucus, since no specific effort was made to exclude it. DNA was extracted from the samples using the MOBIO PowerPlant DNA Isolation Kit following the suggested modifications in Sunagawa et al. (2010). Briefly, approximately 50 mg aliquots of the tissue slurry from each sample were processed with the addition of a lysozyme step and additional, smaller beads to expedite physical lysis. Three extractions were done per coral sample (for a total of 36 extractions) and then recombined by sample after elution of the DNA from the spin column (resulting in 12 DNA samples, one per coral). The DNA samples were quantified with a Thermo Fisher Scientific Quanti-iT PicoGreen dsDNA Assay Kit, per the manufacturer’s protocol.

DNA samples were amplified with primers targeting the V4-V5 hypervariable region (563F/926R) of the 16S rRNA gene: forward primer (5′ AYTGGGYDTAAAGNG) and reverse primer (5′ CCGTCAATTYYTTTRAGTTT). The forward primer was tagged with one of four multiplex (MID) tags so the samples could be combined for sequencing on three plates. Amplification, pooling and 454 sequencing using GS FLX Titanium chemistry were performed by EnGenCore LLC. Sequence data from all samples were deposited in the NCBI SRA under Bioproject number PRJNA305617.

Sequence data were analyzed using the bioinformatic package Quantitative Insights Into Microbial Ecology (QIIME) version 1.8 (Caporaso et al., 2010, Nature Methods 7:335-336, doi:10.1038/nmeth.f.303). Please refer to the file entitled "Lophelia_workflow_2016_10_19.txt," which is included as a supplemental file and details the scripts run in QIIME. The workflow file contains the default or chosen settings used for each script, as well as the names of the input/output files associated with each script.

Keywords section of metadata optimized by correcting variations of theme keyword thesauri and updating/adding keywords.

Added keywords section with USGS persistent identifier as theme keyword.