Bio-Identification of Flake
Samples Using DNA Barcoding
By Lina Martinez, Jeremy Tallman, Richard Moloney,
Chantall Smith, Joanna Wisniewski , Wathsala Kumari
Katherine Te
DNA
BARCODING
•Provides a faster method of accuracy
determining a species when compared
with morphological identification.
•It allows for how species are assembled
within a biological systems .
•Determine the composition/purity of
biological samples .
•The DNA Barcode used here is a section of
the Cytochrome c
oxidase 1, a region involved in the electron phase of
mitochondrial respiration.
 Our group decided to determine if DNA Barcoding
could be used to identify which species of shark
are sold in Australia as “flake”.
 Hypothesis – that different species of shark are
sold under the banner of flake
Primer sequences used in the experiment were:

“A fragment from the 5’ end of COI (~ 648 base pairs) is isolated (the entire gene has~
1500 base pairs).”
COI barcoding (fish)
5’-TGTAAAACGACGGCCAGTCAACCAACCACAAAGACATTGGCAC -3’
α
FORWARD PRIMER – VF2_t1
5’-TGTAAAACGACGGCCAGTCGACTAATCATAAAGATATCGGCAC-3’
α
FORWARD PRIMER – Fish F2_t1
5’-CAGGAAACAGCTATGACACTTCAGGGTGACCGAAGAATCAGAA-3’
α
REVERSE PRIMER – Fish R2_t1
5’-CAGGAAACAGCTATGACACCTCAGGGTGTCCGAARAAYCARAA-3’
α
α
α
REVERSE PRIMER – FR1
For other materials and methods refer to Project Manual (Diploma In Laboratory
Techniques MSL50109)
We had DNA of
of about 580 base pairs
which was the expected
size.

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CONSENSUS SEQUENCES

FLAKE SAMPLE - Galeorhinus galeus

TGGGACAGCCAGGATCTCTTTTAGGAGATGACCAGATTTATAATGTGATCGTAAC
CGCCCATGCTTTTGTAATAATCTTTTTTATGGTTATACCAATCATGATTGGTGGCT
TTGGAAATTGACTAGTTCCATTAATAATTGGTGCGCCTGATATAGCCTTCCCACG
GATAAATAACATAAGCTTCTGACTTCTTCCACCATCATTCCTTCTTCTCCTAGCTT
CTGCCGGAGTAGAAGCTGGAGCAGGTACTGGTTGAACAGTATATCCTCCACTAG
CAAGCAATTTAGCCCATGCTGGACCATCTGTAGATTTAGCCATTTTCTCCCTTCA
TTTAGCCGGTATCTCATCAATCCTAGCCTCAATTAATTTTATTACAACCATTATTA
ACATAAAACCCCCAGCTATTTCCCAATATCAAACACCATTATTTGTTTGATCAATT
CTTGTAACTACTATTCTTCTTCTCCTCTCTCTCCCAGTTCTCGCAGCAGGAATCAC
AATATTACTTACAGACCGTAACCTTAATACCACATTCTTTGACCCTGCAGGTGGA
GGAGACCCAATCCTTTACCAACATTTATTCTGATTCTTC
SAMPLE NUMBER
COLLECTION LOCATION
NAME OF SPECIES
Flake 1
Fish shop (fresh)
Mustelus lenticulatus- Spotted
Estuary Smooth-Hound
Flake 2
Fish shop (cooked)
Flake 3
Supermarket (fresh)
Flake 4
Market (fresh)
Mustelus antarcticus – Gummy
Shark
Galeorhinus galeus- School
Shark, Tope Shark, Soupfin
Shark or Snapper Shark
Mustelus antarcticus – Gummy
Shark
Flake 5
Market (fresh)
Flake 6
Market (fresh)
Flake 7
Supermarket (fresh)
Table 1. Results of DNA sequencing
No Results
No Results
Galeorhinus galeus- School
Shark, Tope Shark, Soupfin
Shark or Snapper Shark
“Just as a unique pattern of bars in a universal product
code (UPC) identifies each item for sale in a store, a DNA
barcode is a DNA sequence that uniquely identifies each
species of living thing” 1
DNA barcoding has been successful in identifying the
species of shark samples used in our experiment.
7 samples of Flake were purchased from different
locations in Melbourne. Genomic DNA was extracted, and
used for PCR. 2 of the 7 samples had a low quality result
and were removed from the data set. 5 were sufficient
enough for analysis.
•Mustelus antarcticus, is a shark in the
family triakidae . It is a slender, grey shark
with white spots along the body and flat,
plate-like teeth for crushing its prey.
•It has a maximum length of between 157 cm
(male) and 175 cm (female). It is found in the
waters around southern Australia,
from Shark Bay in WA to Port Stephens in
NSW to a depth of 350 m.
•Marketed as "flake”.
GUMMY SHARK
http://www.oceanwideimages.com/images/10801/large/gum
my-shark-24M2642-01D.jpg
• Mustelus lenticulatus, is
a houndshark of the family
triakidae, found on the continental
shelves and in estuaries around
southern Australia and New
zealand .
•Its length is up to 125 cm (male)
and 151 cm (female).
•“FISH AND CHIP” = FLAKE
SPOTTED ESTUARY
SMOOTH-HOUND
http://www.southern-edge.com/rig-shark-or-lemonfish/
• Galeorhinus galeus, is
a hound shark of
the family Triakidae.
• Found worldwide in
subtropical seas at depths
of up to 550 metres
(1,800 ft).
• It grows to 2 metres
(6 ft 7 in) long.
SCHOOL SHARK, TOPE
SHARK, SOUPFIN
SHARK OR SNAPPER SHARK,
http://www.exploreuw.com/Galeorhinus_galeus.d
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Our results support our hypothesis. We found 3 different
species are sold as “flake”. This shows that DNA Barcoding
can be used to identify seafood species sold in Australia.
2 out of the 5 shark DNA sequences was that of the
endangered Tope Shark.
The Tope Shark is found on the Red List of the IUCN - The
World Conservation Union.
DNA barcoding can be used to help in the conservation of
endangered species.

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
The government has a National Plan of Action
for the Conservation and Management of
Sharks 2012 – Shark Plan 2
It aims on working on conservation and
managing Australian Fisheries.
Our data suggests that DNA barcoding could
be a good method to assist this plan
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For future experiments a bigger sample size would
help to discover how wide spread endangered species
are being consumed and labelled as “shark”.
The larger the sample size of shark being tested, the
more we can have confirmation of our results.
Our results agree with the results of the students from
New York’s Trinity School who did an experiment on
‘mislabelled’ fish.
We found out that DNA Barcoding is an effective
technique and can be use to identify different species of
sharks sold in the supermarket as “flake”.