Lesson 2_1: Sequence Alignments

Assignment 2_1

Beginning with Margret Dayhoff's creation of the Mutation Data Matrix in the late 1960's, computers have been used more and more to help investigation of similarity amongst protein and DNA sequences. Now there are sites that provide HTML and MUD mediated interactive programs for data submission and analysis. There is even a 1 year old Internet course on Biocomputing. This assignment is meant to familiarize you with theory and facilities for sequence alignments and similarity searches. Please make use of the many links to additional materials in the background reading. There is also some useful information in the GCG Users Guide at the Biotec Center.

1. Read the background information (and also make use of this excellent VSNS site, and for this section, especially the chapter on weighted matrixes; and or check out the tutorials listed below at the Pittsburg Supercomputing Center).
2. Conduct the canned exercise to see what is meant by a multiple sequence alignment.
   • Retrieve seqdata.txt. This file contains three sequences in the FASTA format. Use the select and copy to clip-board functions to paste sequence from this window to the clipboard.
   • Return to this page.
   • Click on the link to an internet version of Clustal W..
   • Enter the sequences. Use copy / paste to aid in the data entry.
   • Check the ClustalW option (it is the default, but just be sure), and press the submit button.
   • You should see a return of the alignment and data submitted, but it might take a few minutes. Note that for large runs, or if you don't want to wait, there is an Email option for receiving the results.
   • Save the alignment report to a local file.
   • Post a link to your saved copy of the HTML report from your home page to complete your assignment.
3. Using a local  copy of ClustalW (save this zip file on your hard drive, rename it to clust.exe, and execute it to unzip the program files; UPDATE - ClustalW has been updated  for a Windowed interface as ClustalX (perhaps more upto date, local copy placed 9/98) on the same set of sequences. As far as I can tell, the two programs are very similar, but the windows version shows a plot of alignment quality, and there may be some options for alignment parameters that differ between the two programs (scoring matrices available, etc). Note the different options that you have, and be sure to read about them [use the help feature and go to the various menu screens to see what options are available; or read the author's comments (ClustalW, ClustalX) (or other helpful descriptions of ClustalW)] so that you understand what the choices mean. Conduct an alignment of sequences that you care about. Post a brief description of why you chose those sequences, and use the report to copy / paste the alignment preformatted text so that you may discuss the significance of conserved regions.

Potentially Useful links:

• Pedro's Molecular Research Tools -A super list of molecular biology tools - you must check this out!
• ClustalW, ClustalX  - ClustalV was (as of 3/96) updated  to ClustalW to provide more rigorous alignment options (using optimal PAM values for example), and to deal intelligently with variable gap penalties (see the author's comments). Here is a second detailed description of ClustalW, together with Pileup, both used for making multiple alignments. ClustalX appears to be a windowed version of ClustalW - see authors comments.
• BCM Search Launcher: Multiple Sequence Alignments - ClustalW and other programs via form page.
• VSNS Course on Biocomputation

Some Interesting Links found at the Pittsburg Supercomputing Center (PSC)

Software available at the PSC

Nucleic Acid and Protein Sequence Analysis software

Sequence Analysis Tutorials

• Introduction to the GCG Wisconsin package - note that after a two month trial that is made available to those attending the PSC training workshops, one has to pay for access to GCG at PSC (money goes to the company selling GCG, not the PSC). There are other alternatives:
  • use a locally provided GCG
  • use the WEB to do what GCG can do
  • use PSC alternative programs
• Sequence Analysis Tutorials Reformatting Sequences
• Primer on Finding Homologous Proteins or Genes
• Sequnce Analysis: Which scoring method should I use?
• A Tutorial on Searching Sequence Databases and Sequence Scoring Methods
• Sequence Analysis Tutorials Multiple Sequence Alignment and Patterns

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