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Before looking at the alignment in Jalview, look at the text results.

Which 2 sequences align with the highest pairwise alignment score?

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Look at the text display of the alignments, shown just below the scores. Try to determine the regions where the 6 sequences align most closely.Many people find this type of text display hard to interpret (a fact that is good to remember when putting together a figure for a presentation or anarticle). At the bottom of the text display is a cladogram. A cladogram is a branched phylogenetic-type tree where the branches are of equal length.Cladograms can show common ancestry, but because the branches have equal lengths, they do not provide an accurate indication of the evolutionarydistance between the branches.

Does the cladogram group together all the human CFTR sequences in the same branch?

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Now, look at the alignment in Jal view. Is it easier to identify the regions where all6 sequences align most closely in the Jalview display?

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Leave both of these windows open (closing the ClustalW text results browser page may cause the Jalview editor pageto disappear automatically). Notice the file number assigned this Jalview sequence alignment. This is to help distinguish it for purposes of comparison from the nextand last alignment in this tutorial, where the gap penalty will be manipulated from the default values.

Finally, repeat the CFTR alignment using the sequences above, but this time change "Gap Open" to 100.(Click on the term "Gap Open" to view the default value.) This will increase the penalty for opening a gap. For more information on gaps and gap penalties,view the EBI help page entitled "About Gaps" . Compare the group alignment scores from the previous alignment with these groupalignment scores. The group alignment scores can be found in the .output file (listed in the results section) under the section which starts:

Start of Multiple Alignment There are 5 groupsAligning...

Does changing the gap open penalty affect the group alignment scores?

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Are the group scores higher or lower when the penalty is increased to 100?

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View the Jalview display of the new alignment. Compare the Jalview displays of the first alignment,and the new alignment with the increased gap penalty, in the regions numbered 1220 to 1250.

Describe the differences in the two alignments in this region, both in terms of the gaps, and in terms of the similarity bargraph displayed below the sequences.

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A look at the ClustalW submission page is enough to see that there are many more parameters that can be manipulated by the user. However, the default values have been optimized and generally should be left alone, except in cases where the user has a definitive justification for making the change. It does frequently occur in biology, though, that the user knows some empirical information that conflicts with the results given by an alignment performed with the default values. In these cases, a scientific argument can be made for altering the parameters to force the alignment to reflect the empirical information.

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Source:  OpenStax, Bios 533 bioinformatics. OpenStax CNX. Sep 24, 2008 Download for free at http://cnx.org/content/col10152/1.16
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