SYSTEMATICS
AP Biology
The implications of our understanding of the mechanisms that create change in organisms – selection, drift, etc - are clear: organisms are related, and so they share common ancestry. It follows that the more closely related two organisms are the more similarities they share.
Systematics is the branch of biology that works to organize all the known life forms into categories that reflect shared genetic histories. In 1735 Carl Linneaus’ proposed a system for grouping, ranking and naming organism based on morphological homologies. His taxonomy classifies organisms into progressively more exclusive groups; the two most specific levels of the hierarchy – genus and species – are used to form the scientific name of the organism.
Today, molecular homologies are used to deduce evolutionary lineages . Such a phylogeny outlines relationships between organisms, an improvement over the grouping of taxonomy.
Phylogeny includes diagrams called cladograms . A cladogram sequences a set of organisms into increasingly smaller groups that possess the traits of the groups before them, as well as features distinct to the new group.
A. CLASS PHYLOGENTETIC TREE
A phylogenetic tree depicts in graphic form the divergence of a species from its ancestors over time. In this model, each member of the lab group will start with the same ancestral DNA sequence. Over time, groups and individuals will undergo random and unique mutations to the
DNA resulting in cumulative differences that can be compared to determine relatedness. Those with the greatest differences diverged the greatest number of years ago. Those with the fewest differences are the most closely related.
1. Everyone in the class shares a common ancestor, for which we have the DNA sequence.
Divide the class in 2. Record this divergence on the whiteboard.
2. Determine how many base changes everyone in Group 1 will make by rolling the dice and adding two to the result. Draw that many numbers from the ‘nucleotide position’ bag to determine where the changes happen in the sequence. Finally, draw a nucleotide letter for each of the numbers to determine what the change will be. Return all numbers & letters to the bags.
3. Diverge from the ancestor again; that is, each group divides into smaller groups again.
Repeat the mutation process for each group, even if the ‘group’ has only one member.
This is your species.
4. Diagram the series of branches the divergences created on the white board, writing the name of a student only at the end – not the base - of each branch.
5. Use the class phylogenetic tree to make a hypothesis about who you are most and least closely related to.

6. Test your hypothesis by counting the number of similarities in DNA between you and multiple other students. Record in the matrix, Table 1.
7. Generate a scatter plot of your matrix data: % similaritiy vs number of shared ancestors.
B. PRIMATE CLADOGRAM: Morphological basis
Linnean taxonomy groups together in the order Primate those mammals with an opposable thumb and/or toe, allowing for a grasping action.
8. Have a discussion about the distinct traits of each of the primates based on the PPT images provided. Discuss how the species might be related based on morphology. Create a hypothesis, in the form of a cladogram, that reflects your predicted.
C. PRIMATE CLADOGRAM: Gene sequence basis
Entrez is a web portal that allows users to search for gene information, such as nucleotide sequence, in databases at the National Center for Biotechnology Information.
BLAST is a program that compares a selected sequence to other sequences in the database and calculates the statistical significance of matches. BLAST can be used to infer functional and evolutionary relationships between sequences as well as help identify members of gene families.
9. Find a gene sequence via ENTREZ
Search a different gene than your lab partners; share data. http://www.ncbi.nlm.nih.gov/gene
At the HOME page, toggle to ‘gene’. Enter the name of a protein, prefaced by ‘human’, into the search bar. Scroll through the results list; choose one/click on one from the list provided.
Suggestions: human insulin, myosin, actin, growth factor, oxytocin, ubiquitin, hemoglobin, myoglobin – try the ‘list of proteins’ at Wikipedia for more ideas. http://en.wikipedia.org/wiki/List_of_proteins
10. Scroll a few pages down to “NCBI Reference sequence” > mRNA + Protein(s) > click on the NM number > (opens new page)
Click ‘run BLAST’ if it is an option in the right-hand bar
If not, Click on “FASTA” > (sequence appears)
Highlight and copy the sequence to import into BLAST at this link: http://blast.ncbi.nlm.nih.gov/Blast.cgi?CMD=Web&PAGE_TYPE=BlastHome

11. Find similar sequences via BLAST
Choose ‘nucleotide BLAST’
Paste in the Entrez sequence if no nm number shows in the field
Check “Search set: Others”
Check “Program Selection: highly similar sequences”
Scroll down and click the BLAST button
Record the value from the ‘max score’ column into the table below
D. RESULTS & ANALYSIS
1. For each protein, calculate the percent similarity of each species compared to the human; record in Table II.
2. Considering the information collected by each member of the lab group, create a cladogram of these primates. If you are unsure about any of the relationships, create variations of your tree to acknowledge those uncertainties. Summarize the relationships of the primates on the tree.

E. DISCUSS
1.
Differentiate between phylogeny and taxonomy. Relate to evolution.
2.
Use the toy animals and a whiteboard to create a taxonomic scheme, and a cladogram.
3.
What do the individual letters in the ancestral DNA of part A represent? What do the changes represent?
4.
Species X and Y have 25 amino acid differences. Species Z and X have 10 amino acid differences in the same protein. Species Y and Z have 27 amino acid differences. a.
Organize the data into a differences matrix. b.
Which organism - X, Y or Z - diverged from the common ancestor first? Which pair of organisms is likely to share more characteristics than the other pairs?
Explain your reasoning. c.
Create a phylogenetic tree showing the evolutionary relationship between X, Y and Z and their common ancestor.
5.
Did your BLAST results identify any non-primates who have a highly similar sequence to your protein? What can you infer about a gene/protein that has a high degree of similarity with non-primate mammals?
6.
Where are genus & species found in the taxonomic hierarchy? What is the relationship between them?
7.
Felis concolor is called Puma, Cougar, & Mountain Lion, depending on your locale.
Why is the scientific name more valuable to a biologist than the common name?

Table I : Similarities Matrix
Students
6
5
3
2
4
1 2 3 4
Table II: Nucleotide similarity
Protein
Homo sapiens
Pan troglodytes
Scientific & common name
Pan paniscus
Gorilla gorilla
Pongo abelli
#
%
5
Nomascus leucogenys
Macaca mulatta

Teacher notes:

The BLAST algorithm calculates similarity scores for local alignments (i.e., the most similar regions between 2 sequences) between the query sequence and subject sequences using specific scoring matrices, and returns a table of the best matches (“hits”) from the database. The hit table includes several useful pieces of information, including the similarity score, query coverage (percent of the query sequence that overlaps the subject sequence), E-value (see below), and max identity (percent similarity between the query and subject sequences over the length of the coverage area). The E (Expect) value of the hit is a parameter that describes the number of hits one can ‘expect’ to see by chance when searching a database of a particular size. Essentially, the E value describes the random background noise.
• Try this alternate alignment tool, which will create a tree from a set of sequences. See example; not sure how to correctly upload the sequences. http://www.phylogeny.fr/
D. Frustrated? Try these LINKS to GENE SEQUENCES:
Homo sapiens myoglobin (MB), transcript variant 1, mRNA http://www.ncbi.nlm.nih.gov/nuccore/NM_005368.2
GNA12 guanine nucleotide binding protein (G protein) alpha 12 [ Homo sapiens ] (actin) http://www.ncbi.nlm.nih.gov/gene/2768
HBB hemoglobin , beta http://www.ncbi.nlm.nih.gov/nuccore/NM_000518.4
KRT18 keratin 18 [ Homo sapiens ] http://www.ncbi.nlm.nih.gov/gene/3875
MYO9B myosin IXB [ Homo sapiens ] http://www.ncbi.nlm.nih.gov/gene/4650
WWP1 WW domain containing E3 ubiquitin protein ligase 1 [ Homo sapiens ] http://www.ncbi.nlm.nih.gov/nuccore/NM_007013.3
Consider using Entrez to find the sequences of other human genes; Use BLAST with these and add the search results to your table

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Eucalyptus does not include species name or a map, yet it is included in the Pacific Coast Tree
Finder - why?
The final step in identifying the Blue Oak is geographic - Quercus douglasii is found in northern
CA and Q. englemanni is native to southern CA. Present an argument for their reclassification as a single species.
Trees to flag at keiser park – go across sports fields and road at south end of park.
Valley oak, Quercus lobata – deep lobes, no points
Blue Oak, Q. douglassii – no lobes, small points,
CA black oak, Q. kelloggii – deep lobes, distinct points
OR ash, Fraxinus latifolia – riparian, 7 compound leaflets/leaf
? Common chokecherry, Prunus virginiana – riparian, watch poison oak!
DEMo in parking lot – coast redwood
Trees from the ‘hood Trees in the south parking lot
Genus common name
Genus
Quercus Oak Prunus common name
Plum
Betula Birch Platanus Sycamore

Morus
Liquidamber
Mul-berry
Sweet-gum
Crataegus
Liriodendro n
Hawthorn
Yellow
Poplar