Part I: Build a phylogenetic tree In this assignment you will be guided to build a build an evolutionary tree using the rcbL gene sequence, which is commonly used to study  the evolutionary relationships between plants,

  • Your answers should be in red so that they can be easily distinguished from the question
  • Hyperlinks should remain blue à confirm that they are functional
  • Any sequences should be pasted on font courier new, size 8-10

 
Part I: Build a phylogenetic tree
In this assignment you will be guided to build a build an evolutionary tree using the rcbL gene sequence, which is commonly used to study  the evolutionary relationships between plants,

  • You will gather data pertaining to the rcbL gene.
  • You will acquire the rbcL gene data from different plant species and upload into MEGAX
  • You will trim and align the sequences
  • You will build a phylogenetic tree
  • You will analyze the phylogenetic tree and draw conclusions about the evolutionary relatedness of the species.

 Do some research to answer the following questions, cite your sources.

    1. What function does the protein product of the rcbL gene have in the plant?
    2. Which organelle does the rbcL gene originate from?
    3. Why the rcbL gene is used to establish phylogeny among plant species?
    4. Why rbcL gene is used for plant barcoding?
    5. What barcoding means?

 

  1. Retrieve the sequences below:
    1. What database can you use to retrieve the sequences?
    2. What format should you use to retrieve the sequences?
    3. What type of sequences are in this data set?
    4. Paste sequences in proper format in the word document?
  2. pen sequences in MEGAX
  3. Edit sequences names to show scientific name only.
  4. Complete the table below:
Common Name Accession # Scientific name
Corn AKQ78206.1
Thale cress QDK58872.1
Rice BAA00147.1
Tobacco CAA77361.1
Potato ABB90049.1
Liverwort AZU95178.1
Sunflower QCX31658.1
Grape ABE47542.1
Cucumber AUZ97435.1
Spinach CAB88737.1
Tomato ABC56308.1
Cilantro AAA84137.1
Pinus AAS78807.1
Oak QHF18935.1
Euglena AEY70827.1

 

  1. Align sequences in MEGAX
  2. Align sequences using Muscle or ClustalW Explain your choice?
  3. Trim the sequences: trim the beginning & ends of the sequences so that they line up nicely and do not contain too much excessive data or gaps
  4. Save the alignment in MEGA file
  5. Paste the beginning and end of the alignment in the word document

 

  1. Use the aligned file to build a phylogenic tree in MEGAX
  2. Statistical Method = Neighbor-Joining
    Test of Phylogeny = Bootstrap
    No. of Bootstrap Replications = 1000 to obtain stable estimates of reliability of the tree.
    Substitution Type start by selecting Nucleotide followed by selecting the Jukes-Cantor Model.
  3. Obtain the Condensed Tree

Note – for question 5 substitution : amino acid and poisson model.
 

  1. Inspect the Condensed Tree and identify the outgroup
  2. Identify the outgroup and explain your reasoning for choosing the outgroup
  3. Right click on the branch that contains your selected outgroup and select from submenu the Place Root option.
  4. Paste the rooted phylogenetic tree

 

  1. Analyze the phylogenetic tree and answer the following questions:
  2. Which species are most closely related?
  3. Give an example of sister taxa.
  4. Why do you think these two taxa are closely related?
  5. How many base pairs were included in your analysis?
  6. How do you think the length of the gene sequence used affects the validity/reliability of your results?
  7. Is there any other species that cluster with the outgroup? If so explain why?

 

  1. Use the aligned file in question 4 to build a phylogenic tree in MEGAX
  2. Statistical Method = Maximum likelihood
    Test of Phylogeny = Bootstrap
    No. of Bootstrap Replications = 1000 to obtain stable estimates of reliability of the tree.
    Obtain the Condensed Tree
  3. Inspect tree and select outgroup
  4. Paste the rooted phylogenetic tree
  5. Analyze the tree and answer the questions in Q#7.
  6. Which tree [Neighbor-Joining or Maximum likelihood] illustrates best the evolutionary relationships among the plant taxa in the rbcL gene data set. Explain in detail.

 PART II: Compare evolution of orthologues

Mitogen-activated protein kinases

The proteins encoded by these genes are members of the MAP kinase family. MAP kinases, also known as extracellular signal-regulated kinases (ERKs), act as an integration point for multiple biochemical signals, and are involved in a wide variety of cellular processes such as cell proliferation, differentiation, gene expression and development. MAP kinases are found in eukaryotes only, but they are fairly diverse and encountered in all animals, fungi and plants, and even in an array of unicellular eukaryotes. The activation of a kinase requires its phosphorylation by upstream kinases. Upon activation, this kinase translocates to the nucleus of the stimulated cells, where it phosphorylates nuclear targets.
In this part of the assignment you will compare the evolution of different MAPK orthologues.
References

  1. NCBI Gene, Gene ID: 132, http://www.ncbi.nlm.nih.gov/gene/132, 12/8/2012
  2. Pearson G, Robinson F, Beers Gibson T, Xu BE, Karandikar M, Berman K, Cobb MH (April 2001). “Mitogen-activated protein (MAP) kinase pathways: regulation and physiological functions”. Endocr. Rev. 22 (2): 153 83. doi:10.1210/er.22.2.153. PMID 11294822

 

  1. Below, paste in FASTA format the MAPK sequences that will be used for the rest of the questions
  2. You will use a total of 15 amino acid sequences
  3. The first 10 are provided
  4. For the remaining 5, you will select any orthologs of your choice
  5. List the source organism (scientific and common name) and accession no hyperlink
  6. Hint 1: if you want a more complete phylogenetic study, you are allowed to include more than 15 sequences
  7. Hint 2: Edit definition line as shown in classà the first 10 characters in the definition line need to be unique
  8. Hint 3: Use underscores to link the definition line terms that you will like to appear on your alignment, i.e., Homo_M15; Danio_M15, Danio_M6, etc.
  9. Hint 4: Select sequences of fairly similar lengths (+ or – ~200 amino acids are ok)
  10. Hint 5: You can use any source/database to find your selected sequences (UniprotKb, Ensembl, etc.)

 

Required Sequences Protein/
Gene Name
Source Organism Scientific Name Common name
(if known)
Length (a.a) Accession no.
Hyperlink
Need to make hyperlinks for required accession no. sequences
Sequences required by instructor
1 MAPK15 Homo sapiens Human 544 Q8TD08
2 MAPK15 Canis familiaris Dog 559 E2RLC0
3 MAPK15 Mus musculus mouse 549 Q80Y86
4 MAPK15 Danio rerio Zebra fish 524 A0A0R4IIR5
5 MAPK15 Xenopus tropicalis Western Clawed frog 593 F6U845
6 MAPK4 Homo sapiens Human 587 P31152
7 MAPK4 Canis familiaris Dog 584 E2RIY9
8 MAPK4 Mus musculus mouse 583 Q6P5G0
9 MAPK6 Homo sapiens Human 721 Q16659
10 MAPK6 Canis familiaris Dog 723 E2QVF3
Sequences selected by student
11    
12      
13    
14    
15      

 

  • Paste all 15 Sequences– Edit definition lines to be shorter yet informative

 
10. Prepare a ClustalW alignment using ClustalOmega and paste it below in full Clustal format  11. Trim sequences = Delete gaps
On the ClustalW alignment below, determine what is the beginning and the end of your alignment should be.
·   On the ClustalW alignment above (Q#10) highlight in yellow or using boxes the areas that you decide will be the beginning and the end of your final alignment.  ·   Sequences outside of this boundary need to be deleted- BUT you will do this directly on MEGAX (on the next section  ·   Hint 1: Find areas of conservation at the beginning and at the end of the alignment and use those to begin and end your alignment  12. Re-do the alignment on MEGAX but use MUSCLE as your algorithm. Take a screen capture of the beginning and the end of your alignment  Beginning: these screen captures are provided as an example. Delete them and replace with yours.              End: these screen captures are provided as an example. Delete them and replace with yours. 13. Edit your alignment a.    Edit the beginning of your MUSCLE alignment by deleting non-conserved areas at the BEGINNING of your alignment. Re-do alignment and paste new screen capture of the beginning of your alignment  Edited Beginning screen capture: b.    Edit the end of your MUSCLE alignment by deleting non-conserved areas at the END of your alignment. Re-do alignment and paste new screen capture of the ending of your alignment Hint: Save this alignment so that you can use it to build your trees. 14. Paste your alignment on ClustalW format below a.    Hint 1: on MEGA Alignment Explorer, go to DATA and select Export Alignment, FASTA Formatb.    Hint 2: Open alignment on Word or Notepad, paste into Clustal Omega window, and select Output format as ClustalW and paste alignment in FULL clustalW format belowc.    Using the shapes below (edit their size as needed), find and box-in at least 2 distinct areas on your alignment have been shown to evolve the least during time (in blue) and those that have evolved the most (in green).

       
c.    Explain well your selection on “b”.  Why did you select these areas?  Explain the possible importance of those areas for protein function for both blue and green area. EXPLAIN IN DETAIL YOUR ANSWER.·   Poor or too short answers will not receive full credit ·   You should use citations found in peer-reviewed scientific publications to support your answer.

  1. Using your MUSCLE ALIGNMENT, build 4 phylogenetic trees WITH bootstrap (1000 trees) using the algorithms listed below and paste the BOOTSTRAP Consensus Tree
    Hint: Test of Phylogeny needs to be Bootstrap for all (1000)

 

  1. Maximum Likelihood Tree from MUSCLE Alignment
  2. Neighbor Joining Tree derived from MUSCLE Alignment
  3. UPGMA Tree derived from MUSCLE Alignment
  4. Maximum Parsimony Tree derived from MUSCLE Alignment

16. Analyze the trees and alignments you constructed and answer the following questions. EXPLAIN IN DETAIL YOUR ANSWER·   Poor or too short answers will not receive full credit ·   You should use citations found in peer-reviewed scientific publications to support your answer.

  1. After analyzing all trees, are there groups of organisms who’s MAPK seem to follow a similar evolutionary pattern? Do they cluster by a specific group of organisms or by type of kinase? Explain your findings and use specific examples from the trees to illustrate your findings
  2. Which one of your phylogenetic groups seemed to have the least branching support? Find evidence in the literature to explain and substantiate your answer
  3. If you were to design a drug that would be use for a condition directly related to Human MAPK16, which orthologue(s) would you consider using and why?

 
Order Now

Calculate a fair price for your paper

Such a cheap price for your free time and healthy sleep

1650 words
-
-
Place an order within a couple of minutes.
Get guaranteed assistance and 100% confidentiality.
Total price: $78
WeCreativez WhatsApp Support
Our customer support team is here to answer your questions. Ask us anything!
👋 Hi, how can I help?