Clustalx program

Gibson , Desmond G. Higgins , 3 and Julie D. Thompson 4, a. Desmond G. Julie D. Author information Article notes Copyright and License information Disclaimer. This article has been cited by other articles in PMC. Abstract The Clustal series of programs are widely used in molecular biology for the multiple alignment of both nucleic acid and protein sequences and for preparing phylogenetic trees.

Table 1. Open in a separate window. Figure 1. Figure 2. Table 2. A comparison of execution times. Higgins D. Gene , 73 , — Myers E. Feng D. Taylor W. Wilbur W. Natl Acad. USA , 80 , — Sneath P. Methods Mol. Saitou N. Thompson J. Compagne F. Bioinformatics , 16 , — Maddison D. Benner S. Protein Eng. Bateman A. Howe K. Bioinformatics , 11 , — In practice, you should use a large number of bootstrap replicates is recommended, even if it means running the program for an hour on a slow computer.

You can also supply a seed number for the random number generator here. Different runs with the same seed will give the same answer. See the documentation for more details. With this option, any alignment positions where ANY of the sequences have a gap will be ignored.

This means that 'like' will be compared to 'like' in all distances, which is highly desirable. It also automatically throws away the most ambiguous parts of the alignment, which are concentrated around gaps usually. The disadvantage is that you may throw away much of the data if there are many gaps which is why it is difficult for us to make it the default. For small divergence say Where possible, this option should be used.

However, for VERY divergent sequences, the distances cannot be reliably corrected. You will be warned if this happens. Even if none of the distances in a data set exceed the reliable threshold, if you bootstrap the data, some of the bootstrap distances may randomly exceed the safe limit.

Three different formats are allowed. None of these displays the tree visually. This format is verbose and lists all of the distances between the sequences and the number of alignment positions used for each.

The tree is described at the end of the file. It lists the sequences that are joined at each alignment step and the branch lengths. After two sequences are joined, it is referred to later as a NODE. This format is the New Hampshire format, used by many phylogenetic analysis packages. It consists of a series of nested parentheses, describing the branching order, with the sequence names and branch lengths. This is the same format used during multiple alignment for the guide trees. The format is described fully in: Maddison, D.

Swofford and W. NEXUS: an extensible file format for systematic information. Systematic Biology The toggle allows them to be placed on the nodes, which is incorrect, but some display packages e.

TreeTool, TreeView and Phylowin only support node labelling but not branch labelling. Care should be taken to note which branches and labels go together.

Clustal X provides a versatile coloring scheme for the sequence alignment display. The sequences or profiles are colored automatically, when they are loaded. Sequences can be colored either by assigning a color to specific residues, or on the basis of an alignment consensus. In the latter case, the alignment consensus is calculated automatically, and the residues in each column are colored according to the consensus character assigned to that column. In this way, you can choose to highlight, for example, conserved hydrophylic or hydrophobic positions in the alignment.

Clustal X automatically looks for a file called 'colprot. By default, if no color parameter file is found, protein sequences are colored by residue as follows:. It can be switched off to show residues as a colored character on a white background. The Color option looks first for the color parameter file as described above and, if no file is found, uses the default residue-specific colors.

The format of the color parameter file is described below. The first section is optional and is identified by the header rgbindex. If this section exists, each color used in the file must be named and the rgb values specified on a scale from 0 to 1.

If the rgb index section is not found, the following set of hard-coded colors will be used. RED 0. The second section is optional and is identified by the header consensus. It defines how the consensus is calculated.

The third section is identified by the header color, and defines how colors are assigned to each residue in the alignment.

Clustal X provides an indication of the quality of an alignment by plotting a 'conservation score' for each column of the alignment. A high score indicates a well-conserved column; a low score indicates low conservation.

The quality curve is drawn below the alignment. Two methods are also provided to indicate single residues or sequence segments which score badly in the alignment. Low-scoring residues are expected to occur at a moderate frequency in all the sequences because of their steady divergence due to the natural processes of evolution.

The most divergent sequences are likely to have the most outliers. However, the highlighted residues are especially useful in pointing to sequence misalignments. Note that clustering of highlighted residues is a strong indication of misalignment.

This can arise due to various reasons, for example:. Partial or total misalignments caused by a failure in the alignment algorithm.

Usually only in difficult alignment cases. Partial or total misalignments because at least one of the sequences in the given set is partly or completely unrelated to the other sequences. It is up to the user to check that the set of sequences are alignable. Frameshift translation errors in a protein sequence causing local mismatched regions to be heavily highlighted. These are surprisingly common in database entries.

If suspected, a 3-frame translation of the source DNA needs to be examined. Occasionally, highlighted residues may point to regions of some biological significance. This might happen for example if a protein alignment contains a sequence which has acquired new functions relative to the main sequence set. It is important to exclude other explanations, such as error or the natural divergence of sequences, before invoking a biological explanation.

Unreliable regions in the alignment can be highlighted using the Low-Scoring Segments option. A sequence-weighted profile is used to indicate any segments in the sequences which score badly. The segment display can then be toggled on or off without having to repeat the time-consuming calculations.

Increase the scale to display more segments; decrease the scale to remove the least significant. The matrix is used to calculate the sequence- weighted profile scores. A more stringent matrix which only gives a high score to identities and the most favoured conservative substitutions, may be more suitable when the sequences are closely related. This option automatically recalculates the low-scoring segments. The previous system used by ClustalW, in which matches score 1.

A new matrix can be read from a file on disk, if the filename consists only of lower case characters. You can customise the column 'quality scores' plotted underneath the alignment display using the following options. For more information about the weight matrices, see the help above for the Low-scoring Segments Weight Matrix. The low-scoring segment display can be toggled on or off.

This option does not recalculate the profile scores. This option highlights individual residues which score badly in the alignment quality calculations. Residues which score exceptionally low are highlighted by using a white character on a grey background.

The quality scores that are plotted underneath the alignment display can also be saved in a text file. Each column in the alignment is written on one line in the output file, with the value of the quality score at the end of the line. Only the sequences currently selected in the display are written to the file. One use for quality scores is to color residues in a protein structure by sequence conservation.

In this way conserved surface residues can be highlighted to locate functional regions such as ligand-binding sites. A11 A12 A A1n A21 A22 A Am1 Am2 Am We also have a residue comparison matrix of size R where C i,j is the score for aligning residue i with residue j. To do this, we define an R-dimensional sequence space. For the jth position in the alignment, each sequence consists of a single residue which is assigned a point S in the space.

S has R dimensions, and for sequence i, the rth dimension is defined as:. We then calculate a consensus value for the jth position in the alignment. This value X also has R dimensions, and the rth dimension is defined as:. Now we can calculate the distance Di between each sequence i and the consensus position X in the R-dimensional space. The quality score for the jth position in the alignment is defined as the mean of the sequence distances Di.

If you want to report a bug please make sure, that you are using the most recent version of Clustal. Please also check if the bug has already been reported on our Bugzilla webpage. We would like to thank all Clustal users who have send in feedback, bug reports and feature requests.

Recent work on Clustal was mainly funded by Science Foundation Ireland. Webservers You don't necessarily have to go through the hassle to install Clustal on your computer. Clustal W and Clustal X version 2. Bioinformatics , 23, Multiple sequence alignment with the Clustal series of programs.

Nucleic Acids Res. Multiple sequence alignment with Clustal X. Trends Biochem Sci.