Biology – The TreeBiology
of Life
The Tree of Life
Mapping Evolutionary History
S
MATERIALS
scissors
yarn, approx. 2 m
cientists understand that over long periods of time, DNA and amino acid
sequences in organisms can change. As the number of differences between
DNA sequences increase, species begin to diverge from one another. Given
enough time, new species can emerge from the same ancestor. Scientists trying
to piece together segments of evolutionary history can do this by looking at the
number of shared traits between species. A large number of shared traits are
considered strong evidence that species are closely related.
Although scientists originally grouped organisms using physical traits, today
they are able to group organisms more accurately using amino acid or nucleotide
sequences coded in the genes. These groupings are used to create branched
diagrams called cladograms. Much like a family tree, cladograms are scientists’
attempt to accurately map out evolutionary history.
Molecular cladograms are created by selecting genes or sequences of amino acids
that are shared by the organisms under study. Scientists then search for “updates”
in the patterns, features known as characters, that appear in some of the species
but are not in the common ancestor. Organisms with these updated characters are
considered to be more recent arrivals on Earth. Because variations to the original
nucleotide sequence resulted from gene mutations, they provide clues to how
different organisms diverge from shared ancestors. Comparing patterns of these
various characters changing over time allows scientists to reconstruct a likely
evolutionary history.
When creating cladograms, a branching tree-like diagram must be constructed.
The names of the organisms are placed at the top of the lines (e.g., A, B, and C).
Shared features are placed in solid boxes along the branches, and the common
ancestor is placed in a circle at the base of the cladogram.
Assume a character will evolve only once, so if different organisms display that
character they should be placed into groups closer to one another. In other words,
the more similar two organisms are the closer their evolutionary relationship
and the closer they will be on the cladogram. This also means the two organisms
shared a common ancestor more recently than other organisms under study. Two
examples of cladogram styles are shown in Figure 1.
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Biology – The Tree of Life
Figure 1. Cladograms
Once biologists could view organisms at the molecular level, they quickly
determined physical traits did not provide accurate evolutionary maps.
Sequencing the genome of various organisms showed changes as small as one
nucleotide could indicate even the most subtle of differences between species at
each sequenced gene site. Biologists began collecting information in an attempt to
piece together the evolutionary pathways. One important comparison involves the
genes for cytochrome c.
Cytochrome c codes for a protein attached to the inner mitochondrial membrane
of eukaryotes. This protein is an electron carrier in oxidative phosphorylation of
cellular respiration. It moves electrons through the membrane toward the final
electron acceptor. Because most organisms have cytochrome c, protein sequence
variations are often used to determine phylogenic relationships.
This activity compares sequence variations found in cytochrome c subunit 1
for several types of dolphins, whales, and other organisms and then uses the
information to depict evolutionary history using a cladogram.
PURPOSE
This lab will use genomic information to compare the cytochrome c protein
sequences of several organisms. Thousands of gene sequences for various
organisms are stored in a database with the National Center for Biotechnology
Information (NCBI). Although there are several ways to retrieve sequencing
information from this database, this activity will reference the organism’s
scientific name.
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Biology – The Tree of Life
PROCEDURE
PART I
Use the information in Table 1 to create a cladogram. Remember to box the
distinguishing character along the branching lines. As each organism is singled
out from the others, ensure its name is placed at the end of each branch.
Table 1. Common Characteristics
Organism
Hagfish
Shark
Toad
Snake
Spiny
anteater
Horse
Heart with
Chambers
Jaws
Bony
Skeleton
Amniotic
Egg
Hair
Placental
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
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+
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Biology – The Tree of Life
PROCEDURE (CONTINUED)
PART II
1. Obtain a bag with pictures and the common names of several species of
whales, dolphins, and other organisms.
2. Organize the pictures into groups based upon shared physical traits. Use the
yarn provided to create a possible cladogram with these organisms based
upon your groupings.
3. Create a data table with a list of the organisms in each group and the
characteristics used to place these animals into the same group.
4. Find your Pre-Lab Exercises. Look at the cladogram you created and
compare it to the scientific names of each species. Identify species that share
the same genus but may have been placed into different groups referencing
only the individual’s appearance. Create a second data table with new groups
based on your pre-lab assignment.
5. Highlight any animals that changed from the data table in Step 3. List the
organism(s) that were grouped incorrectly and provide a reason for your
original placement.
6. Using the new information, redesign the cladogram making any changes that
may show more correct evolutionary relationships. Once this is finalized,
make a sketch of this cladogram.
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Biology – The Tree of Life
PROCEDURE (CONTINUED)
PART III
1. Obtain an NCBI BLAST reference sheet for cytochrome c for the group of
animals in Part II. This analysis compares the protein sequence of sample
organisms to that of the blue whale’s cytochrome c sequence.
2. Look at this BLAST reference sheet. The first animal listed is the blue
whale, identified by its scientific name, Balaenoptera musculus. This sample
compares cytochrome c from a blue whale to itself, which means there will
be a 100% match.
Compare the percent identity match of the other species to the blue whale.
As differences appear in the sequences, the percent similarity will decline. In
other words, the species are increasingly different in their protein sequence
for cytochrome c.
3. Create a data table to record some of this information. The data table should
include space for the common and scientific names of each animal. Also
include a column to record the percent identity, the identities fraction and
finally, the sequence differences.
The identity fraction shows a ratio of the number of protein sequence
matches compared to the total number of sequences for cytochrome c. As
an example, when the BLAST reference sheet lists “Identities = 425/512,”
it means there are a total of 512 sequences in this protein and the compared
species had 425 matches to the blue whale.
Calculate the sequence differences (512 – 425), which is 87 variation sites
for this example. Remember, as the number of differences increases the more
distantly related two species must be.
4. Complete the data table for each animal in the chart.
5. Create a cladogram using the pictures and yarn showing the most likely
evolutionary history of these 12 organisms based upon the cytochrome c
sequences. Once completed, sketch the final version into the lab report.
Remember, the more the sequences are alike the closer the organisms should
be on the evolutionary tree.
6. Clean up the area and return all items to the plastic bag.
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Biology – The Tree of Life
PRE-LAB EXERCISES
Find the complete scientific classification (kingdom to species) for the organisms
listed in Table 2.
Common Name
Amazon river dolphin
Blue whale
Bottlenose dolphin
Bullhead shark
California gray whale
Common porpoise
Finback whale
Hippopotamus
Humpback whale
Killer whale
Long-beaked common dolphin
Polar bear
Spotted dolphin
White-beaked dolphin
Kingdom
Class
Order
Table 2. List of Organisms
Phylum
Family
Genus
Species
6
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Biology – The Tree of Life
ANALYSIS
PART I
Create a cladogram in the space provided to show the evolutionary relationships
among the organisms in Table 1.
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
PART II
1. Create a cladogram in the space provided to show the evolutionary
relationships among the organisms in Table 2.
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
2. Create a data table with the animals organized into groups and the
characteristics used to put the organism into that group.
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
3. Reorganize your previous data table based on shared physical traits and
scientific names.
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
4. Highlight any animals that have changed groups between your two tables.
List the organism(s) that were grouped incorrectly and provide a reason for
your original placement.
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
5. Create a cladogram of the organisms using your recreated data table
(referencing their scientific names).
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
PART III
1. Create a data table to record the information from the NCBI BLAST
reference sheet:
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Biology – The Tree of Life
ANALYSIS (CONTINUED)
2. Create a cladogram that represents a likely evolutionary history of the
organisms based on the cytochrome c sequenced genome.
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Biology – The Tree of Life
CONCLUSION QUESTIONS
1. Cladograms convey important information to scientists about a species under
study. List four different things the cladogram from Part I conveys about the
species involved.
2. List some of the challenges in creating a cladogram based only on physical
evidence.
3. What is cytochrome c?
4. Why were the sequence variations in cytochrome c useful in determining
evolutionary divergence among the animals in this study?
5. How did your cladogram based on physical traits (morphology) compare to
the cladogram based on the cytochrome c evidence?
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Biology – The Tree of Life
CONCLUSION QUESTIONS (CONTINUED)
6. What evidence is more accurate for determining evolutionary changes,
morphology or molecular sequencing? Justify your claim.
7. View the cytochrome c cladograms created by other groups. What are some
reasons these cladograms may be different even though each was developed
using the same sequencing information?
8. How do cladograms support the idea that evolution drives the diversity and
unity of life?
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Biology – The Tree of Life
EXTENSION: MOLECULAR CLOCK
Scientists estimate the blue whale diverged from the fin whale approximately
10.3 million years ago. Assuming mutations will occur at the same DNA location
at approximately the same rate throughout time, calculate the mutation rate for the
cytochrome c protein per 1 million years. Show your work.
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Biology – The Tree of Life
ITEM 1 – NCBI REFERENCE SEQUENCE FOR CYTOCHROME C OXIDASE SUBUNIT I (1/2)
NCBI BLAST: COX1 - REFERENCE SEQUENCE: NC _001601.1
Gene ID: 807733
updated 19-Sep-2012
Gene symbol
COX1
Gene type
protein coding
RefSeq status
REVIEWED
Organism
Balaenoptera musculus
Lineage:
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
Mammalia; Eutheria; Laurasiatheria; Cetartiodactyla; Cetacea;
GENE ID: 807733 COX1 | cytochrome c oxidase subunit I [Balaenoptera musculus]
Score = 1030 bits (2662), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 516/516 (100%), Positives = 516/516 (100%), Gaps = 0/516 (0%)
GENE ID: 807613 COX1 | cytochrome c oxidase subunit I [Balaenoptera physalus]
Score = 1025 bits (2662), Expect = 0.0, Method: Compositional matrix adjust.
C O N S U M A B L E
Mysticeti; Balaenopteridae; Balaenoptera
GENE ID: 3337172 COX1 | cytochrome c oxidase subunit I [Megaptera novaeangliae]
Score = 1022 bits (2639), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 511/516 (98%), Positives = 514/516 (99%), Gaps = 0/516 (0%)
L E S S O N
Identities = 513/516 (99%), Positives = 515/516 (99%), Gaps = 0/516 (0%)
GENE ID: 2658485 COX1 | cytochrome c oxidase subunit I [Eschrichtius robustus]
Score = 1021 bits (2639), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 508/516 (98%), Positives = 514/516 (99%), Gaps = 0/516 (0%)
GENE ID: 2658614 COX1 | cytochrome c oxidase subunit I [Lagenorhynchus albirostris]
Score = 986 bits (2548), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 499/516 (97%), Positives = 509/516 (99%), Gaps = 0/516 (0%)
GENE ID: 9978248 COX1 | cytochrome c oxidase subunit I [Orcinus orca]
Score = 968 bits (2503), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 495/516 (96%), Positives = 506/516 (98%), Gaps = 0/516 (0%)
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Biology – The Tree of Life
ITEM 2 – NCBI REFERENCE SEQUENCE FOR CYTOCHROME C OXIDASE SUBUNIT I (2/2)
GENE ID: 7411782 COX1 | cytochrome c oxidase subunit I [Delphinus capensis]
Score = 968 bits (2503), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 500/516 (97%), Positives = 509/516 (99%), Gaps = 0/516 (0%)
GENE ID: 7412033 COX1 | cytochrome c oxidase subunit I [Stenella attenuata]
Score = 968 bits (2502), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 499/516 (97%), Positives = 509/516 (99%), Gaps = 0/516 (0%)
GENE ID: 2658388 COX1 | cytochrome c oxidase subunit I [Inia geoffrensis]
GENE ID: 7412047 COX1 | cytochrome c oxidase subunit I [Tursiops truncatus]
Score = 964 bits (2493), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 499/516 (97%), Positives = 508/516 (98%), Gaps = 0/516 (0%)
GENE ID: 2658445 COX1 | cytochrome c oxidase subunit I [Phocoena phocoena]
Score = 962 bits (2486), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 492/514 (96%), Positives = 507/514 (99%), Gaps = 0/514 (0%)
GENE ID: 804864 COX1 | cytochrome c oxidase subunit I [Ursus maritimus]
L E S S O N
Identities = 490/516 (95%), Positives = 507/516 (98%), Gaps = 0/516 (0%)
C O N S U M A B L E
Score = 967 bits (2499), Expect = 0.0, Method: Compositional matrix adjust.
Score = 941 bits (2431), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 485/514 (94%), Positives = 505/514 (98%), Gaps = 0/514 (0%)
GENE ID: 808675 COX1 | cytochrome c oxidase subunit I [Hippopotamus amphibius]
Score = 952 bits (2461), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 493/512 (96%), Positives = 508/512 (99%), Gaps = 0/512 (0%)
GENE ID: 3283879 COX1 | cytochrome c oxidase subunit I [Heterodontus francisco]
Score = 827 bits (2136), Expect = 0.0, Method: Compositional matrix adjust.
Identities = 449/514 (87%), Positives = 490/514 (95%), Gaps = 0/514 (0%)
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Biology – The Tree of Life
ITEM 3 – VARIOUS ORGANISMS (1/3)
White-beaked dolphin
Common porpoise
Spotted dolphin
Bottlenose dolphin
Long-beaked common dolphin
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L E S S O N
C O N S U M A B L E
Amazon river dolphin
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Amazon river dolphin
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Delphinidae
Genus: Lagenorhynchus
Species: albirostris
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Iniidae
Genus: Inia
Species: geoffrensis
Spotted dolphin
Common porpoise
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Delphinidae
Genus: Stenella
Species: attenuata
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Phocoenidae
Genus: Phocoena
Species: phocoena
Long-beaked common dolphin
Bottlenose dolphin
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Delphinidae
Genus: Delphinus
Species: capensis
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Delphinidae
Genus: Tursiops
Species: truncatus
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L E S S O N
White-beaked dolphin
C O N S U M A B L E
Biology – The Tree of Life
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Biology – The Tree of Life
ITEM 4 – VARIOUS ORGANISMS (2/3)
Killer whale
Finback whale
California gray whale
Humpback whale
Bullhead shark
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L E S S O N
C O N S U M A B L E
Blue whale
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Blue whale
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Delphinidae
Genus: Orcinus
Species: orca
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Balaenopteridae
Genus: Balaenoptera
Species: musculus
California gray whale
Finback whale
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Eschrichtiidae
Genus: Eschrichtius
Species: robustus
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Balaenopteridae
Genus: Balaenoptera
Species: physalus
Bullhead shark
Humpback whale
Kingdom: Animalia
Phylum: Chordata
Class: Chondrichthyes
Order: Heterodontiformes
Family: Heterodontidae
Genus: Heterodontus
Species: francisco
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Cetacea
Family: Balaenopteridae
Genus: Megaptera
Species: novaeangliae
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L E S S O N
Killer whale
C O N S U M A B L E
Biology – The Tree of Life
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Biology – The Tree of Life
ITEM 5 – VARIOUS ORGANISMS (3/3)
Polar bear
L E S S O N
C O N S U M A B L E
Hippopotamus
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Biology – The Tree of Life
Hippopotamus
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Carnivora
Family: Ursidae
Genus: Ursus
Species: martimus
Kingdom: Animalia
Phylum: Chordata
Class: Mammalia
Order: Artiodactyla
Family: Hippopotamidae
Genus: Hippopotamus
Species: amphibius
L E S S O N
C O N S U M A B L E
Polar bear
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