Multiple Sequence Alignment Tools


What is T-Coffee?

T-Coffee is a multiple sequence alignment package. You can use T-Coffee to align sequences or to combine the output of your favorite alignment methods (Clustal, Mafft, Probcons, Muscle...) into one unique alignment (M-Coffee).

T-Coffee can align Protein, DNA and RNA sequences. It is also able to combine sequence information with protein structural information (3D-Coffee/Expresso), profile information (PSI-Coffee) or RNA secondary structures (R-Coffee).

This presentation gives an overview of the T-Coffee algorithm and of the original implementation and validatation of the package. The full list of associated papers is available on the following links: algorithms and servers. The original T-Coffee paper is:

T-Coffee: A novel method for multiple sequence alignments.
Notredame,Higgins,Heringa,JMB,302(205-217)2000 [pdf][medline]

T-Coffee is a freeware opensource package distributed under the GNU public license



T-Coffee runs on UNIX and UNIX-like platforms (Linux, cygwin and MacOSX).Check out the Source Code from Github or Download the latest Binaries as provided below.

Note that if you want to use the New Regressive T-Coffee allowing Large scale alignments, we recommand Downloading the Beta Version below. A brief documentation is available .

Stable Version: [ Stable ]
Beta Version: [ Beta ]
Nature Protocols Version: [ click here ]
Previous Versions: [ Archives ]
Older Versions: cedric.notredame@gmail.com


T-Coffee is distributed as pre-compiled binaries for Linux and Mac OSX platforms with a guided installation procedure. This is the smoothest and quickest way to install T-Coffee on your machine, since it comes with all required components and does not need any special user account.

Linux/Unix users

  1. download the installer package available here (or the latest beta here);
  2. open a terminal window, move to the download path, and grant execute permission to the installer typing the following command:
    chmod +x T-COFFEE_installer_<version>.bin
  3. launch the installer
  4. when the installation procedure has finished open a new terminal window (so that changes made by the installer are effective) and type the following command to verify your installation:
    t_coffee -version

Mac OSX users

  1. download the compressed package named T-COFFEE_installer_<version>.dmg available here (or the latest beta here);
  2. click to open the download file and launch the contained executeble installer;
  3. when the installation procedure has finisched open a new terminal window (so that changes made by the installer are effective) and type the following command to verify your installation:
    t_coffee -version

Windows users

T-Coffee pre-compiled installer is not available for Windows operating system. Windows users need to compiled and install it using the source distribution package as described below.

An easier alternative could be to install a Linux guest on your Windows workstation using a using a common virtualization tool (like VMware, VirtualBox, etc) and install T-Coffee in the virtualized machine.

Compile and install from sources

Advanced users and anyone requiring custom compilation or installation can download the T-Coffee source distribution packages from Github and follow the procedure to compile source code available here.


mode Regressive t_coffee -seq=foo.seq -reg
Seq. All
Accuracy High - Suitable for very large datasets
mode Fast t_coffee foo.seq
Seq. All
Accuracy Medium
mode Accurate t_coffee foo.seq -mode accurate
Seq. Proteins
Accuracy High but slow
mode R-Coffee t_coffee foo.seq -mode rcoffee
Seq. RNA
Accuracy High
mode Very Fast t_coffee foo.seq -mode quickaln
Seq. All
Accuracy Medium Low
mode Low Mem. t_coffee foo.seq -mode memory
Seq. RNA
Accuracy Medium
mode Expresso t_coffee foo.seq -mode expresso
Seq. All
Accuracy High
mode M-Coffee t_coffee foo.seq -mode mcoffee
Seq. All
Accuracy Medium High


MANUAL [html][pdf][epub]
Nature Protocols


  • Q: What is the most accurate mode?

    A: for proteins, try t_coffee yourseq -mode accurate. For RNA, try: t_coffee -mode rcoffee.

  • Q: What is the meaning of the color coded html output?

    A: Red bits are very reliable, while green and blue bits are unreliable. This color scheme is named the TCS index and it uses the consistency among pairwise alignments for this estimationto estimate reliability. An extensive validation of the TCS is available from the original MBE paper and more details are given on the TCS homepage. Online evaluations can be carried out on the TCS server.

  • Q: Which mode should I use on www.tcoffee.org server to get the best alignment ?

    A: If you have proteins, Expresso is likely to be the most accurate flavor, as it uses potential known 3D-Structures similar to your sequences. Otherwise, our benchmarks indicate that M-Coffee is the next best alternative. M-Coffee runs most known Multiple Sequence Alignment methods and combines them into one final alignment. The most accurate mode (-mode=accurate) is now available at this T-Coffee web server mirror

  • Q: Which Paper Shall I cite ?

    A: It is very important you cite us if you use our ressources. We depend on it for funding! If you use the local version of T-Coffee, cite the original 2000 paper (cf above). Otherwise, cite the paper corresponding to the server you have been using (click on the "cite" button associated with every server on www.tcoffee.org).

  • Q: What to do if I run out of memory ? (I)

    A: Here is the less memory hungry mode of t_coffee (and slightly less accurate):

    t_coffee  your_seq.pep -mode memory  

  • Q: What to do if I still run out of memory ? (II)

    A: Divide your sequences in subgroups, align them individually and use

    t_coffee -profile aln1 aln2 aln3

  • Q: What is the meaning of the name "T-Coffee" ?

    A: T-Coffee stands for Tree based Consistency Objective Function For AlignmEnt Evaluation.


  • T-Coffee Google Group
  • T-Coffee Github developers community
  • MARNA: RNA multiple Structural Alignment
  • T-Coffee is incorporated in both and Biopython.
  • T-Coffee and M-Coffee are part of the Debian distribution ( A big thanks to Charles Plessy).
  • T-Coffee and M-Coffee are part of the Ubuntu distribution.
  • T-Coffee is integrated in the MacVector sequence analys tool.


    T-Coffee relies on your feeback. Please send me an Email if you wish to make a request, a comment, or report a bug!

    Dr. Cedric Notredame, PhD.
    Group Leader
    Comparative Bioinformatics Group
    Bioinformatics and Genomics Programme
    Center for Genomic Regulation (CRG)
    Dr Aiguader, 88
    08003 Barcelona
    Email: cedric.notredame@gmail.com
    HOME : http://www.tcoffee.org/index.html
    Phone: +34 933 160 271