Barcoding for CoML:
Assessing Zooplankton Diversity
Rob Jennings – University of Connecticut, USA
Photos by R.R. Hopcroft – University of Alaska, USA
L.P. Madin – Woods Hole Oceanographic Institution, USA
N. Copley – Woods Hole Oceanographic Institution, USA
CMarZ Steering Group Meeting
6 November – Ocean Research Institute, University of Tokyo
CoML Ocean Realm Field Projects
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Natural Geography in Shore Areas – NaGISA
Gulf of Maine Area Program – GOMA
Pacific Ocean Shelf Tracking – POST
Census of Diversity of Abyssal Marine Life – CeDAMar
Tagging of Pacific Pelagics – TOPP
Patterns and Processes of Ecosystems in the Northern
Mid-Atlantic – MAR-ECO
Biogeography of Chemosynthetic Ecosystems – ChEss
Arctic Ocean Diversity – ArcOD
International Census of Marine Microbes – ICOMM
 Census of Marine Zooplankton - CMarZ
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Global Census of Marine Life on Seamounts – CenSeam
Global Census of Coral Reef Ecosystems – CREEFS
Continental Margin Ecosystems on a Worldwide Scale
– CoMargE
Census of Antarctic Marine Life – CAML
What is DNA Barcoding?
 Definition: Derivation of short DNA sequence(s) that
enables species identification or recognition in a
particular domain of life (e.g., eucaryotes).
 Focus to date: For animals, a 658 base-pair fragment
of the mitochondrial gene, cytochrome oxidase
subunit I (mtCOI).
 Barcode of Life Initiative (BOLI) will resolve
barcodes for named species and use a barcoding
approach to assess undescribed biological diversity.
What isn’t DNA Barcoding?
 It is not intended to, in any way, supplant or
invalidate existing taxonomic practice.
 It is not DNA taxonomy; it does not equate species
identity, formally or informally, with a particular DNA
sequence.
 It is not intended to duplicate or compete with
efforts to resolve deep phylogeny (e.g., Assembling
the Tree of Life, ATOL).
Why Barcode Zooplankton?
 DNA is particularly useful to study animal
plankton, because the organisms are frequently
rare, fragile, and/or small.
 Evolutionarily-conserved body plans for some
groups (e.g., copepods) makes morphological
identification difficult and mistakes likely.
 Many species are widespread or circumglobal;
DNA can be used to evaluate taxonomic
significance of geographic variation.
 DNA-based species identification will speed
analysis of samples for known species.
 Zooplankton will test barcode protocols, since
15 animal groups (phyla) are represented.
Barcoding Goals for CMarZ
 Link morphological / molecular systematic analysis
for global zooplankton assemblage
 DNA barcode ~7,000 described species in 15 phyla
 Submit DNA, specimen & collection data:
- Barcode section of GenBank
- CMarZ database with environmental data
- Searchable from OBIS portal
 Reveal cryptic species within circumglobal species
by population genetic analysis
 Discover new species by sampling biodiversity
hotspots, unexplored ocean regions, deep sea
 Assess zooplankton diversity by ‘environmental
sequencing’ of unsorted samples
 Develop automatable DNA chip-based approaches
and protocols to identify and quantify species
Barcoding Ostracods (Crustacea)
Ostracod species
differed in mtCOI
sequence by 15% to 38%
for 19 species (15 more
completed).
Barcoding for Ostracods:
• Good species
identification
• Cryptic variation
within species
Barcoding Euphausiids (Crustacea)
Fourteen of 86 euphausiid
species were identified by
Peter Wiebe.
50 euphausiids  including 19
species of Euphausia  have
been barcoded to date.
Barcoding for Euphausiids:
• Good at species identification
• Can reveal cryptic species
Barcoding Planktonic Gastropods
MtCOI barcodes for 17
species of gastropods
(pteropods and
heteropods) differed
by 1.4% to 44%.
Barcoding for
Gastropods:
• Good at species
identification
• Less consistent
species phylogeny
Barcoding Medusozoans (Cnidaria)
• Barcoding is done by Brian
Ortman (UConn/USA).
• 13 species of siphonophores
• greatly expanded, much of
siphonophore diversity (160
spp.)
Barcoding Arctic Zooplankton
 ArcOD (Arctic Ocean Diversity)
sends identified specimens of Arctic
zooplankton for barcoding by
CMarZ.
 Comprehensive DNA database of
~210 species of the Central Arctic
assemblage is underway.
 DNA obtained from ~50 species
already, comprising several dates
and locations; work in progress at
UConn
CMarZ Barcoding Structure
and Operations
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Barcoding cruises
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“DNA sensitive” collection protocolsanyone can collect
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Standardized and web-available sampling
and processing protocols
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Taxonomic training  Molecular training
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Increased ability to use preserved (e.g.
formalin) samples
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Distributed network of barcoding centers
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Local taxonomic expertise
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Care of specimens
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Capacity-building
CMarZ Barcoding Centers
• Existing barcoding centers
• UConn, USA
 Oviedo, Spain
 ORI, Japan
• Emergent barcoding centers
• Qingdao, China
 Goa, India
Results and Conclusions
 Global-scale analysis of zooplankton species
diversity; integrated morphological and
molecular analysis.
 DNA barcodes for all ~6,800 known species of 15
phyla in the zooplankton.
 Species discovery by CMarZ will double number
of zooplankton species
- sampling new ocean regions (deep sea)
- cryptic species within circumglobal species
Got specimens? CMarZ encourages and challenges the CoML community to
provide identified specimens of holozooplankton groups for barcoding. A DNA
barcode (700 base-pair region of mtCOI with replicates / metadata) will be
determined for appropriately preserved (i.e., frozen or in alcohol) and identified
holozooplankton specimens. Contact Rob Jennings (rob.jennings@uconn.edu).
Got data? DNA barcodes for zooplankton should be submitted to:
1. BARCODE section of GenBank (www.ncbi.nlm.nih.gov/BankIt/barcode)
2. Barcode of Life Database (BOLD; see www.barcodinglife.org.
UCONN address for specimen shipment:
Ann Bucklin
Department of Marine Sciences
University of Connecticut - Avery Point
1080 Shennecossett Road, Groton, CT 06340 USA
Email: ann.bucklin@uconn.edu
Acknowledgements
CMarZ Steering Group Members
CoML Zooplankton Workshop
Portsmouth, NH USA, March 2004
Demetrio Boltovskoy (Arg.)
Janet Bradford-Grieve (NZ)
Ann Bucklin (USA)
Colomban de Vargas (France)
Ruben Escribano (Chile)
Steven Haddock (USA)
Steve Hay (UK)
Russell R. Hopcroft (USA)
Ahmet Kideys (Turkey)
Laurence P. Madin (USA)
Webjørn Melle (Norway)
Vijayalakshmi Nair (India)
Shuhei Nishida (Japan)
Mark D. Ohman (USA)
Francesc Pagés (Spain)
Annelies Pierrot-Bults
(Netherlands)
Chris Reid (UK)
Sigrid Schiel (Germany)
Sun Song (China)
Erik Thuesen (USA)
Hans Verheye (South Africa)
Peter Wiebe (USA)
Acknowledgements
UConn Team DNA
Brian Ortman
(Univ. Conn. Ph.D. student)
Leo Blanco Bercial
(Univ. Oviedo, Spain)
Ebru Unal
(Univ. Conn. Ph.D. student)
Paola Batta Lona
(Univ. Conn. Res. Assist.)
Lisa Nigro – not shown
(Univ. Conn. Lab Manager)
CJ Sweetman – not shown
(Univ. Conn. Guest Researcher)
UConn Team DNA on the RV Ron Brown
April 2006