Richard Markham and colleagues (Proc. Natl. Acad. Sci. 95(21):12568-73, 1998), published data on the pattern of HIV evolution and the rate of CD4 T-cell decline sequence in 15 subjects by collecting blood samples at six month intervals for up to four years. From each visit, a 285 base pair region within the env gene from all the varieties of the gene (referred to as clones) were sequenced and CD4 T-cell counts were made. Six hundred and sixty-six nucleotide sequences were deposited in GenBank providing a rich resource for looking closely at the patterns of change in HIV over time.
Exploring HIV Evolution (686 kb pdf)
Number of visits: between 3-9 for each patient
Number of clones observed per subject per visit: 2-18
Total number of sequences available: 666
CD4 cell counts for each visit
Summary table of information available on the subjects studied in Markham et al. (1998).
1 The CD4 count for time 1 is reported in Table 1 of Markham et al. (1998); the other values are estimated from the Figure 1 of the same publication.
2 The paper reports 5 visits for subject 2, only 3 visits were identified in the GenBank records.
HIV-1 GP120 core complex with CD4 and a neutralizing human antibody (link to http://www.rcsb.org and I believe it is ID number 1GC1A)
Analyzing DNA Sequences for HIV env Protein
GETTING INTO BIOLOGY WORKBENCH
1. Open Netscape Navigator or Internet Explorer and type the following URL into the Location box: http://workbench.sdsc.edu
OPENING YOUR ACCOUNT
2. Click on the hotlink "Enter the Biology Workbench 3.2," which is blue and is large and underlined. This will bring up a small screen.
3. Supply your username and password. If you donít have one, click the NEW USER button and create an account for yourself.
4. Click on the "OK" button. This will give you a new screen.
STARTING A NEW SESSION
5. Scroll down and click on the "Session Tools" button.
6. Click on the "NEW" button.
7. At the next screen, create a session name (e.g. "HIV env") in the white box to the right of the words "Session Description."
8. Click on the "Start New Session" button.
UPLOADING NUCLEIC ACID SEQUENCES
9. Click on the "Nucleic Tools" button.
10. Scroll down and click on the "Add" button in the middle of the top line of buttons.
11. Click the "Browse" button.
12. Find the Desktop and double click on the "HIV Sequences" folder. This will open the folder. Go to the popup menu for "Files of type:". Select "All Files" from the menu. This will bring up the data for all subjects. Choose your first subject by double clicking on that subject's number.
13. Click on the "Upload File" button. At this point, you will be able to see the sequences on your screen.
14. IMPORTANT!!!! Click the "SAVE" button at the top of the page.
15. Repeat steps 12-16 to add the data for a second subject.
ALIGNING SELECTED SEQUENCES
16. Click on any number of DNA sequences PER SUBJECT to activate them. A small checkmark should appear in the box to the left of all the sequences that you will want to analyze.
Note: The names of the files will look like the following: S10V5-6. This stands for subject 10, visit 5, clone 6.
17. Click on the "CLUSTALW" button on the right of the second row of buttons at the bottom of the screen. This will give you a new screen with the selected sequences listed.
18. On this screen, click the "Submit" button. Then scroll down to see your alignment.
19. Your alignment can be downloaded to your PC and viewed more easily in GeneDoc.
MAKING AN EVOLUTIONARY TREE
20. Scroll to the top of the page and click on the "Import Alignment(s)" button.
21. In the next screen, click in the box to the left of "CLUSTALW-Nucleic" to activate the set of aligned sequences.
22. Click on the "DRAWGRAM" button.
23. Click on the "Submit" button. Then scroll down to see your tree.24. Print your tree by choosing "Print" under the "File" menu.
Author: Sam Donovan -- 2002