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  • Mitochondrial gene barcoding of NE Atlantic Octocorals

    Abstract

    Mitochondrial gene barcoding is a useful way of identifying unknown specimens; however, their effectiveness in distinguishing between anthozoans remains controversial. It has been demonstrated that anthazoans have unusually slow evolving mitochondrial genomes and therefore lack sufficient variation for differentiation at low taxonomic levels. However, DNA barcodes have proven useful for analysis of scleratinian corals, and studies on the less well known octocorals, a sub-class of anthozoa, have shown some improvements in taxonomic identification with the use of more gene sequences to gain greater phylogenetic resolution. Here we present work on the phylogenetic analysis of octocorals collected throughout the NW Atlantic using three mitochondrial gene barcodes to test the ability of this combination to distinguish between known octocoral species. Octocoral specimens were collected from the Bay of Biscay and waters surrounding the Azores, Ireland, NW Scotland, and Iceland. Total DNA was extracted from samples and each sequenced for three mitochondrial regions: the ND2 subunit of NADH dehydrogenase, the “Folmer region” of COI with an adjacent intergenic region igr1, and the octocoral specific mitochondrial protein coding gene msh1. Phylogenetic reconstruction was performed on 144 samples by combining all three barcode regions and results were compared with morphological identifications of the specimens.

    Introduction

    1. Octocorals are important...
    2. NE Atlantic has lots of octos
    3. DNA barcodes for octos (McFadden work)
    4. recent work on connectivity of cold water corals

    McFadden's coral barcoding references (McFadden 2004) (McFADDEN 2011)

    Seamount endemism (or not) based on octocoral genetics (Thoma 2009), (Baco 2012), (Miller 2010)

    Complete genomes of octocorals (Figueroa 2013)

    Bamboo corals have different arrangement of mtdna (Brugler 2008)

    CO1 isn't great for corals (France 2002)

    General biology of octocorals (Watling 2011)

    Molecular phylogenies (Dolan 2013), (McFadden 2010), (McFadden 2006), (Herrera 2012), (Taylor 2015)

    Methods

    1. Specimen collection

    Collections were made during two cruises in the northeast Atlantic. First in April 2010 using the ROV Holland 1 during cruise CE10014 on the Celtic Explorer (Marine Institute, Ireland). Second in September 2011 using the ROV Victor 6000 during the "BobEco" cruise aboard Pourquoi Pas (Ifremer, France). Additional samples were collected by Fernando Tempera (Azores) and Stefan Ragnarsson (Iceland) and Kirsty Kemp (Scotland) (Wright 2014).

    1. Specimen identifications (MT & ABH)

    Specimens were identified by Michelle Taylor and Andrea Braga-Henriques.

    1. Sample prep

    Subsamples of specimens were stored in 75-95% ethanol. DNA extraction used the DNeasy Tissue kit (QIAGEN Ltd. West Sussex, UK), following the manufacturer’s instructions with the modification of a 48 hour digestion period. PCRs were performed on successful extractions using 20ul reaction volumes consisting of: Xul PCR mastermix (Qiagen ltd), 1ul forward and reverse primers (0.2 pm concentration). Primers used were XXX for mutS, YYY for ND2, and ZZZ for CO1

    1. Analysis

    Results

    We sampled X specimens, representing Y taxa for Z regions...

    Discussion