MB 409 - Midterm #1 - 2/14/00
1. What are the three primary evolutionary branches of life? (5 points)
Bacteria, Archaea, and Eukarya
2. Give an example of a generally-accepted fundamental difference in the molcular biology of prokaryotes and eukaryotes, and explain why it is not such a fundamental difference after all. (5 points)
Eukaryotes have a nucleus and prokaryots do not. But... prokaryotes do have a nuclear region, the nucleoid, distinct from the cytoplasm, where nuclear things happen. Some Bacteria even have a membrane around the nucleod!
3. What is the difference between taxonomy and phylogeny? (5 points)
Taxonomies are artificial classification schemes
Phylogenies are natural, evolutionary relationships
4. Give an example of how all known organisms are the same. (5 points)
All organisms encode information in DNA, using the same universal 3-letter code.
5. How was the "endosymbiont theory" for the origin of mitochondria and chloroplasts proven? (5 points)
By molecular phylogenetic analysis of mitochondrial and chloroplast-encoded ssu-rRNAs.
6. What are the steps of a molecular phylogenetic analysis? (5 points)
7. What are the properties of the small subunit ribosomal RNA that make it well-suited for moecular phylogenetic analyses? (10 points)
In most cases, the gene encoding the RNA in the small subunit of the ribosome (ssu rRNA) is the best choice because:
8. Align these sequences: (5 points)
Sequence A - CACACUUCGGAGAG -CACACUUCGGAGAG- Sequence B - CCCACUUUGUGGG -CCCACUUU-GUGGG- Sequence C - ACACACUUCGGAGAG ACACACUUCGGAGAG- Sequence D - ACCCCCUUUGGGGGGU ACCCCCUUUGGGGGGU
9. Align these RNAs: (5 points)
RNA A RNA B RNA C RNA A -GCGAGCUUUCGGGACGC--- RNA B UCGGA-CCUUU-GGACCG--- RNA C -UUUACAAGAAAUUAAAAGAC
10. You have isolated a new organism from the exploded remnants of a very bad can of Spam. From this culture, you have identified an RNA known as "RNA 42", which is found only in the Thermus/Deinococcus group of the Bacteria. There are only three known genera in this group: Deinococcus, Deinobacter and Thermus. Deinococcus and Deinobacter are very closely related, and are together known as the 'Deinococci'. Below is a signature table for RNA 42, for which the Thermus aquaticus RNA is the reference. To which of these two groups does your organism belong? (10 points - show your work!)
Position Deinococci Thermus UNKNOWN ----------------------------------------------- 3 G OK U X G 10 C OK G/UX C 11 A OK C X A 17 -/UOK A X - 19 C OK U/COK C ----------------------------------------------- 5/5 1/5
Therefore the unknown is a member of the Deinococci.
11. Create a similarity matrix with this alignment. (5 points)
A B C D Sequence A - UUUUUCGAAA - - - - Sequence B - UAUUUCGAUA 0.8 - - - Sequence C - AAUUUCGAUU 0.6 0.8 - - Sequence D - CCCUUCGGGG 0.4 0.4 0.4 -
12. Why does the similarity between two sequences underestimate their evolutionary distance? (5 points)
Because of the occurence of multiple changes at single sites. The frequency of these must be estimated from the observed sequence difference.
e.g. A->G->C is 2 changes, but is counted as a single difference
e.g. A->G->A is 2 changes, but is counted as 0 changes!
13. Answer the following questions about this tree: (2 points each for a total of 10 points)
13a. What are the two most closely related sequences? A and B
13b. What are the two most distantly related sequences? C and F
13c. Which sequence is most closely related to D? A
13d. Which sequence is probably the outgroup? F
13e. Circle the last common ancestor of sequences A, B and C.
14. Convert the phenogram in question 13 to a dendrogram (be careful about distances!). (5 points)
15. Briefly describe a method other than molecular phylogenetic analysis for determining phylogenetic relationships. (5 points)
Example: DNA:DNA hybridization.
The extent that the genomic DNAs of 2 species will hybridize is a general measure of how much sequence similarity there is between the genomes, and therefore how closely related they are. This method is widely used to define bacterial species - in general, two organisms are considered to be the same species if the DNA:DNA hybridization is 70% or greater, or different species of the same genus if they have measurable hybridization less than 70%
16. The tree of below is based on valine tRNA synthetase sequences - the enzyme that charges the appropriate valine-specific tRNAs with valine. Sequences in black are from Bacteria, those in orange are from Archaea, and those in blue are from eukaryotic nucleii. These sequences are highly conserved, and so the alignment of these sequences looks very good. Each of the proteins was tested for valine charging ability (i.e. they are all real valine-specific tRNA synthetases), and each of these organisms contains only a single valine tRNA synthetase. How does this tree differ from the ssu-rRNA "3 Kingdom" tree, and how would you explain this? (The root of the tree was identified using a tRNA synthetase specific for a different amino acid.) (10 points)
Sce = Saccharomyces cerevisiae
Hsa = Homo sapiens
Aae = Aquifex aeolicus
Bsu = Bacillus subtilis
Mge = Mycoplasma genitalium
Mpn = Mycoplasma pneumoniae
Hin = Haemophilus pneumoniae
Eco = Escherichia coli
Mtu = Mycobacterium tuberculosis
Ssp = Synechococcus sp. PCC6803
Ctr = Chlamydia trachomatis
Hpy = Helicobacter pylori
Bbu = Borrelia burgdorferi
Tpa = Treponema pallidum
Afu = Archaeoglobus fulgidus
Mth = Methanobacterium thermoautotrophicum
Mja = Methanococcus jannaschii
Pho = Pyrococcus horikoshi
V = valine-specific tRNA synthetase
In the rRNA-based 3-Kingdom tree, Archaea, Bacteria, and Eukarya form 3 separate lusters. In this tree, the eukaryal sequence fall amongst the bacterial sequences. The most likely expanation for this is that the original valine-specific tRNA synthetase of an ancestral eukaryote was replaced by a bacterial gene acquired by horizontal transfer.