1.      Go to the lab and make a hypothesis by ranking the 8 organisms on the following page in Table 1 by how closely related they are to humans. This part is very important. Please do NOT forget to do it!!!

 

2.      Open a new word document. Type your name and class period at the top.

 

3.      Open Mozilla Firefox (DO NOT USE INTERNET EXPLORER). Go to www.uniprot.org

 

4.      At the bottom of the page under “Core data”, click on “Protein Knowledgebase.”

 

                    

 

5.      Type hbb human in the query box. This is an abbreviation for the protein hemoglobin-B (subunit beta) for humans. Hit “search.”

6.      Click on the little box before the first human’s hemoglobin subunit beta and add it to your basket.

7.      Repeat steps 4 and 5 for the following organisms.

Organisms

 hbb Horse

hbb Sheep

hbb Atlantic Salmon

 hbb Western Painted Turtle

hbb Nile Crocodile

hbb Lowland Gorilla

hbb Port Jackson Shark

 

8.      You should now have eight organisms in your basket. Click on the basket and then the “full view” button. Find the “Print screen” button on the top line of the computer keyboard. This will copy an image of the screen. Paste the image in the word document that you opened.   Crop it so only the results of your basket show.

9.      Select all eight organism by clicking on the little box before each organism and then click on the align button. Please be patient. This may take a few minutes!!!

 

 

 

 

 

 

10.  On the left hand side under “Display”, the “Alignment” is already selected. Select both the “Tree” and the “Result info.”

11.  Scroll to the very bottom of the page and complete Table 1 in the lab by recording the entry number and the scientific name (labeled as “organism”). Remember to Capitalize the Genus and make the species lowercase. Underline each name to indicate that scientific names should be in italics.

 

12.  Scroll up to the middle of the page under the “Tree”. This is a cladogram or phylogenic tree of your organisms. Find the “Print screen” button on the top line of the computer keyboard. This will copy an image of the screen. Paste the image into the same word document.   Crop it!!  Enlarge it if need be.

 

13.  Print this document. Close the word document. Do not close the webpage!!!

14.  Write the common name of the organism next to the entry number on your phylogenetic tree (cladogram). NOTE: They are not always in order. Check the entry numbers and entry names carefully!

 

15.   On the webpage, scroll down to the “Results Information”. This chart shows similarities in amino acid sequences of the hemoglobin-B protein. The letters represent identical amino acids, the dashes indicate many differences between the animals.  Copy this information.

 

16.  Open a new word document.

 

 

17.  Copy and paste all sequences to the new word document. (You cannot use the print screen for this part).  Note that the identification numbers are at the beginning of the sequence. Type the name of the animal next to the identification number to make it easier for you to keep track of what you are doing. NOTE: They are not always in order. Check the species numbers carefully!

 

18.  Go to http://fasta.bioch.virginia.edu/fasta/lalign.htm

Or        http://fasta.bioch.virginia.edu  and click on Local DNA alignments (lalign)

 

 

19.  Copy one sequence (not including identification number) into the first box. Copy another in the second box. Look at Table 2 before you begin. You will be making comparisons between all of the chosen animals, so keep track of what you are doing.

 

Text Box: Each of these boxes has one sequence- do not include the species identification number.

 

 

 

20.  Click on “align sequences.”

 

 

 

21.  Find the set of results of the “147 aa overlap” or “146 aa overlap”. In parentheses you will see the percent similarity between these two organisms in amino acids of hemoglobin B. In the above example the two animals have a 93.2% similarity in their amino acid sequences for this protein.

22.  Record this information in Table 2. Repeat until all remaining comparisons have been made.