Service Retirement

We remind you that it is not long until the EBI's Wise2DBA and Promoterwise services are retired on 15th April 2018. Alternatives can be found at Exonerate, BWA or BLAT. If you have any concerns, please contact us via support.



T-Coffee is a multiple sequence alignment program. The main characteristic of T-Coffee is that it will allow you to combine results obtained with several alignment methods. By default, T-Coffee will compare all your sequences two by two, producing a global alignment and a series of local alignments (using lalign). The program will then combine all these alignments into a multiple alignment.

Official Website
Download Software

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 in GCG, FASTA, EMBL (Nucleotide only), GenBank, PIR, NBRF, PHYLIP or UniProtKB/Swiss-Prot (Protein only) 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 currently a sequence input limit of 500 sequences and 1MB of data.

Sequence File Upload

A file containing three or more valid sequences in any format (GCG, FASTA, EMBL (Nucleotide only), GenBank, PIR, NBRF, PHYLIP or UniProtKB/Swiss-Prot (Protein only)) 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 currently a file upload limit of 500 sequences and 1MB of data.

STEP 2 - Set your Parameters


Matrix series to use when generating the multiple sequence alignment. The program goes through the chosen matrix series, spanning the full range of amino acid distances.

Matrix (Protein Only) Description Abbreviation
None none
BLOSUM (Henikoff) These matrices appear to be the best available for carrying out data base similarity searches. blosum
PAM (Dayhoff) These have been extremely widely used since the late '70s. We use the PAM 350 matrix. pam

Default value is: None [none]


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 3 - 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 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).


T-Coffee: A novel method for fast and accurate multiple sequence alignment.
(2000 September 01) Journal of molecular biology 302 (1) :205-217
The EMBL-EBI bioinformatics web and programmatic tools framework.
(2015 April 06) Nucleic acids research 43 (W1) :W580-4
Analysis Tool Web Services from the EMBL-EBI.
(2013 May 13) Nucleic acids research 41 (Web Server issue) :W597-600