ClustalW2

Introduction

ClustalW2 is a general purpose multiple sequence alignment program for DNA or proteins. It attempts to calculate the best match for the selected sequences, and lines them up so that the identities, similarities and differences can be seen.

FAQs
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How to use this tool

Running a tool from the web form is a simple multiple steps process, starting at the top of the page and following the steps to the bottom.

Each tool has at least 2 steps, but most of them have more:

  • The first steps are usually where the user sets the tool input (e.g. sequences, databases...)
  • In the following steps, the user has the possibility to change the default tool parameters
  • And finally, the last step is always the tool submission step, where the user can specify a title to be associated with the results and an email address for email notification. Using the submit button will effectively submit the information specified previously in the form to launch the tool on the server

Note that the parameters are validated prior to launching the tool on the server and in the event of a missing or wrong combination of parameters, the user will be notified directly in the form.

Step 1 - Sequence

Sequence Input Window

Three or more sequences to be aligned can be entered directly into this form. Sequences can be be in GCG, FASTA, EMBL, PIR, NBRF or UniProtKB/Swiss-Prot format. Partially formatted sequences are not accepted. Adding a return to the end of the sequence may help certain applications understand the input. Note that directly using data from word processors may yield unpredictable results as hidden/control characters may be present. There is a limit of 500 sequences or 1MB of data.

Sequence File Upload

A file containing three or more valid sequences in any format (GCG, FASTA, EMBL, PIR, NBRF or UniProtKB/Swiss-Prot) can be uploaded and used as input for the multiple sequence alignment. Word processor files may yield unpredictable results as hidden/control characters may be present in the files. It is best to save files with the Unix format option to avoid hidden Windows characters. There is a limit of 500 sequences or 1MB of data.

Sequence Type

Indicates if the sequences to align are protein or nucleotide (DNA/RNA).

Type Abbreviation
Protein protein
DNA dna

Default value is: Protein [protein]

Step 2 - Pairwise Alignment Options

Alignment Type

The alignment method used to perform the pairwise alignments used to generate the guide tree.

Output Format Description Abbreviation
slow Slow, but accurate slow
fast Fast, but approximate fast

Default value is: slow

Protein Weight Matrix (PW)

Slow pairwise alignment protein sequence comparison matrix series used to score alignment.

Matrix (Protein Only) Description Abbreviation
BLOSUM blosum
PAM pam
Gonnet gonnet
ID id

Default value is: Gonnet [gonnet]

DNA Weight Matrix (PW)

Slow pairwise alignment nucleotide sequence comparison matrix used to score alignment.

Matrix (Protein Only) Description Abbreviation
IUB iub
ClustalW clustalw

Default value is: IUB [iub]

Gap Open (PW)

Slow pairwise alignment score for the first residue in a gap.

Default value is: 10

Gap Extension (PW)

Slow pairwise alignment score for each additional residue in a gap.

Default value is: 0.1

KTUP

Fast pairwise alignment word size used to find matches between the sequences. Decrease for sensitivity; increase for speed.

Default value is: 1

Window Length

Fast pairwise alignment window size for joining word matches. Decrease for speed; increase for sensitivity.

Default value is: 5

Score Type

Fast pairwise alignment score type to output.

Order Description Abbreviation
percent percent
absolute absolute

Default value is: percent

Top Diags

Fast pairwise alignment number of match regions are used to create the pairwise alignment. Decrease for speed; increase for sensitivity.

Default value is: 5

Pair Gap

Fast pairwise alignment gap penalty for each gap created.

Default value is: 3

Step 3 - Multiple Sequence Alignment Options

Protein Weight Matrix

Multiple alignment protein sequence comparison matrix series used to score the alignment.

Matrix (Protein Only) Description Abbreviation
BLOSUM blosum
PAM pam
Gonnet gonnet
ID id

Default value is: Gonnet [gonnet]

DNA Weight Matrix

Multiple alignment nucleotide sequence comparison matrix used to score the alignment.

Matrix (Protein Only) Description Abbreviation
IUB iub
ClustalW clustalw

Default value is: IUB [iub]

Gap Open

Multiple alignment penalty for the first residue in a gap.

Default value is: 10

Gap Extension

Multiple alignment penalty for each additional residue in a gap.

Default value is: 0.20

Gap Distances

Multiple alignment gaps that are closer together than this distance are penalised.

Default value is: 5

No End Gaps

Multiple alignment disable the gap seperation penalty when scoring gaps the the ends of the alignment

Order Description Abbreviation
no false
yes true

Default value is: no [false]

Iteration

Multiple alignment improvement iteration type

Order Description Abbreviation
none No iteration none
tree Iteration at each step of alignment process tree
alignment Iteration only on final alignment alignment

Default value is: none

Num Iter

Maximum number of iterations to perform

Default value is: 1

Clustering

Clustering type.

Order Description Abbreviation
NJ Neighbour-joining (Saitou and Nei 1987) NJ
UPGMA UPGMA clustering UPGMA

Default value is: NJ

Output

Format for generated multiple sequence alignment.

Order Description Abbreviation
Clustal w/ numbers Clustal alignment format with base/residue numbering aln1
Clustal w/o numbers Clustal alignment format without base/residue numbering aln2
GCG MSF GCG Multiple Sequence File (MSF) alignment format gcg
PHYLIP PHYLIP interleaved alignment format phylip
NEXUS NEXUS alignment format nexus
NBRF/PIR NBRF or PIR sequence format pir
GDE GDE sequence format gde
Pearson/FASTA Pearson or FASTA sequence format fasta

Default value is: Clustal w/ numbers [aln1]

Order

The order in which the sequences appear in the final alignment

Order Description Abbreviation
aligned Determined by the guide tree aligned
input Same order as the input sequences input

Default value is: aligned

Step 4 - Submission

Job title

It's possible to identify the tool result by giving it a name. This name will be associated to the results and might appear in some of the graphical representations of the results.

Email Notification

Running a tool is usually an interactive process, the results are delivered directly to the browser when they become available. Depending on the tool and its input parameters, this may take quite a long time. It's possible to be notified by email when the job is finished by simply ticking the box "Be notified by email". An email with a link to the results will be sent to the email address specified in the corresponding text box. Email notifications require valid email addresses.

Email Address

If email notification is requested, then a valid Internet email address in the form joe@example.org must be provided. This is not required when running the tool interactively (The results will be delivered to the browser window when they are ready).

References

Clustal W and Clustal X version 2.0.
(2007 Nov 01) Bioinformatics (Oxford, England) 23 (21) :2947-8
A new bioinformatics analysis tools framework at EMBL-EBI.
(2010 Jul) Nucleic acids research 38 (Web Server issue) :W695-9
Analysis Tool Web Services from the EMBL-EBI.
(2013 Jul) Nucleic acids research 41 (Web Server issue) :W597-600