Cold-water corals are conspicuous in the waters off Eastern Canada. Despite that, there are few DNA sequence records from specimens collected in the region available in GenBank, and not all species recorded in the region have sequence data regardless of geographic origin. This can limit the use of eDNA techniques to detect and identify corals. Our objective was to sequence and publish sequences for two octocoral DNA barcoding markers: CO1 and MutS. We sequenced and deposited 36 sequences to GenBank from 19 specimens representing three sea pen taxa (Octocorallia: Pennatuloidea): Distichoptilum gracile, Pennatula aculeata, and Protoptilum carpenteri. Identification of all specimens was confirmed by B. M. Neves before submission. Specimens and DNA tissues were donated to the Canadian Museum of Nature, where they are currently stored. This publication is part 1 of a series of GenBank submissions by our lab.
Specimens were collected from across the Northwest Atlantic and originate from depths ranging between 200-1924 meters. Specimens were collected as part of research vessel multispecies trawl surveys or remotely operated vehicle (ROV ROPOS) surveys. DNA was isolated and purified using the QIAgen DNeasy Blood and Tissue kit, with an initial overnight incubation with Proteinase K. Two commonly used octocoral barcoding regions were amplified using previously described primers: 1) COII8068F (McFadden et al., 2004) and COIOCTR (France and Hoover, 2002) for the CO1 gene, and 2) ND42599F (France and Hoover, 2002) and mut3458R (Sánchez et al., 2003) for the MutS gene. Amplifications were conducted using 12.5 µl of Green DreamTaq Master Mix (Thermo Fisher Scientific), 1 µl of template DNA, 0.5 µl of each 10 µM forward and reverse primers, 0.5 µl of 10 µM reverse primer, and 10.5 µl of water. Thermocycling was run as follows: 3 min of initial denaturation at 95 °C, followed by 40 cycles at 95 °C for 30 s, 30 s at annealing temperature of 48 °C, then 65 s at an extension temperature of 72 °C, and a final elongation at 72 °C for 4 min. PCR products were cleaned using Agencourt AMPure XP Beads (Beckman Coulter) and sent to The Center for Advanced Genomics, Toronto, Canada for Sanger sequencing. Sequences were visualized and aligned using Geneious Prime 2022.0.2. Obtained sequences have been deposited in GenBank under accession numbers OQ569768- OQ569784 and OQ420359- OQ420377.
This work was funded by Fisheries and Oceans Canada under an Enhanced Regional Capacity grant (2020-2021) and the Marine Conservation Targets (MCT) program (2021-2024), Newfoundland and Labrador Region.