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Technical |
Centre National De LA
Recherche scientifique (France)
CeNTRO De REGULACIO GENOMICA (SPAIN)
Cédric Notredame
www.tcoffee.org
T-Coffee:
Technical Documentation
T-Coffee Technical Documentation
(Version 8.96, July 2010)
www.tcoffee.org
T-Coffee, seq_reformat
PSI-Coffee, 3D-Coffee, Expresso, M-Coffee,
R-Coffee, APDB, iRMSD, T-RMSD
ã
Cédric Notredame, Centro de Regulacio Genomica, Centre National de la Recherche
Scientifique, France
T-Coffee is distributed under the Gnu Public License................................................................. 6
T-Coffee code can be re-used freely............................................................................................. 6
T-Coffee can be incorporated in most pipelines:
Plug-in/Plug-out…......................................... 6
Contributors................................................................................................................................... 7
Addresses....................................................................................................................................... 7
T-Coffee.......................................................................................................................................... 8
Mocca............................................................................................................................................ 10
CORE............................................................................................................................................. 10
Other Contributions.................................................................................................................... 10
Bug Reports and Feedback......................................................................................................... 10
Third Party Packages and On Demand Installations................................................................ 11
Standard Installation of T-Coffee............................................................................................... 11
Automated
Installation......................................................................................................................................................................... 11
Mac-OsX........................................................................................................................................................................................................ 11
Unix................................................................................................................................................................................................................. 11
Unix.................................................................................................................................................................................................................... 12
Microsoft Windows/Cygwin............................................................................................................................................................ 13
MAC osX, Linux........................................................................................................................................................................................... 13
CLUSTER Installation............................................................................................................................................................................. 13
If you have PDB installed:.................................................................................................................................................................... 14
Installing BLAST for T-Coffee...................................................................................................... 14
Why Do I need BLAST with T-Coffee?......................................................................................................................................... 14
Using the EBI BLAST Client................................................................................................................................................................ 15
Using the NCBI BLAST Client............................................................................................................................................................. 15
Using another Client................................................................................................................................................................................ 16
Using a BLAST local version on UNIX.......................................................................................................................................... 16
Using a BLAST local version on Windows/cygwin............................................................................................................ 17
BLAST+.......................................................................................................................................................................................................... 17
ORIGINAL NCBI BLAST.......................................................................................................................................................................... 17
Installing Other Companion Packages....................................................................................... 17
Installation of PSI-Coffee and Expresso..................................................................................... 19
Installation of M-Coffee............................................................................................................... 19
Automated Installation......................................................................................................................................................................... 20
Manual Installation.................................................................................................................................................................................. 20
Installation of APDB and iRMSD.................................................................................................. 22
Installation of tRMSD................................................................................................................... 22
Installation of seq_reformat........................................................................................................ 23
Installation of extract_from_pdb................................................................................................ 23
Installation of 3D-Coffee/Expresso............................................................................................ 23
Automated Installation......................................................................................................................................................................... 23
Manual Installation.................................................................................................................................................................................. 23
Installing Fugue for T-Coffee............................................................................................................................................................. 24
Installation of R-Coffee................................................................................................................ 24
Automated Installation......................................................................................................................................................................... 24
Manual Installation.................................................................................................................................................................................. 25
Installing ProbbonsRNA for R-Coffee.......................................................................................................................................... 25
Installing Consan for R-Coffee.......................................................................................................................................................... 25
T-COFFEE...................................................................................................................................... 26
M-Coffee....................................................................................................................................... 26
Expresso....................................................................................................................................... 27
R-Coffee........................................................................................................................................ 27
iRMSD and APDB.......................................................................................................................... 28
tRMSD........................................................................................................................................... 28
MOCCA.......................................................................................................................................... 29
Environment Variables............................................................................................................... 31
http_proxy_4_TCOFFEE........................................................................................................................................................................ 31
email_4_TCOFFEE..................................................................................................................................................................................... 32
DIR_4_TCOFFEE......................................................................................................................................................................................... 32
TMP_4_TCOFFEE....................................................................................................................................................................................... 32
CACHE_4_TCOFFEE................................................................................................................................................................................. 32
PLUGINS_4_TCOFFEE............................................................................................................................................................................. 32
NO_ERROR_REPORT_4_TCOFFEE.................................................................................................................................................. 32
PDB_DIR.......................................................................................................................................................................................................... 32
NO_WARNING_4_TCOFFEE................................................................................................................................................................ 32
UNIQUE_DIR_4_TCOFFEE.................................................................................................................................................................... 32
Setting up the T-Coffee environment variables........................................................................ 32
Well Behaved Parameters.......................................................................................................... 33
Separation...................................................................................................................................................................................................... 33
Posix.................................................................................................................................................................................................................. 33
Entering the right parameters.......................................................................................................................................................... 33
Parameters Syntax...................................................................................................................... 34
No Flag........................................................................................................................................................................................................... 34
-parameters................................................................................................................................................................................................. 34
-t_coffee_defaults...................................................................................................................................................................................... 35
-mode............................................................................................................................................................................................................. 35
-score [Deprecated]................................................................................................................................................................................ 35
-evaluate....................................................................................................................................................................................................... 35
-convert [cw].............................................................................................................................................................................................. 36
-do_align [cw]............................................................................................................................................................................................ 36
Special Parameters...................................................................................................................... 36
-version......................................................................................................................................................................................................... 36
-proxy............................................................................................................................................................................................................. 36
-email............................................................................................................................................................................................................. 36
-check_configuration.............................................................................................................................................................................. 37
-cache............................................................................................................................................................................................................. 37
-update.......................................................................................................................................................................................................... 37
-full_log......................................................................................................................................................................................................... 37
-plugins......................................................................................................................................................................................................... 37
-other_pg...................................................................................................................................................................................................... 37
Input............................................................................................................................................. 38
Sequence Input........................................................................................................................................................................................... 38
-infile [cw]................................................................................................................................................................................................... 38
-in (Cf –in from the Method and Library Input
section)........................................................................................................ 38
-get_type....................................................................................................................................................................................................... 38
-type [cw]..................................................................................................................................................................................................... 38
-seq.................................................................................................................................................................................................................. 38
-seq_source.................................................................................................................................................................................................. 38
Structure Input.......................................................................................................................................................................................... 39
-pdb................................................................................................................................................................................................................. 39
Tree Input...................................................................................................................................................................................................... 39
-usetree......................................................................................................................................................................................................... 39
Structures, Sequences Methods and Library Input via
the -in Flag......................................................................... 39
-in..................................................................................................................................................................................................................... 40
Profile Input................................................................................................................................................................................................. 41
-profile........................................................................................................................................................................................................... 41
-profile1 [cw]............................................................................................................................................................................................. 42
-profile2 [cw]............................................................................................................................................................................................. 42
Alignment Computation.............................................................................................................. 42
Library Computation: Methods...................................................................................................................................................... 42
-lalign_n_top............................................................................................................................................................................................... 42
-align_pdb_param_file........................................................................................................................................................................... 42
-align_pdb_hasch_mode....................................................................................................................................................................... 42
Library Computation: Extension.................................................................................................................................................... 43
-lib_list [Unsupported].......................................................................................................................................................................... 43
-do_normalise............................................................................................................................................................................................ 43
-extend.......................................................................................................................................................................................................... 43
-extend_mode............................................................................................................................................................................................ 43
-max_n_pair................................................................................................................................................................................................ 44
-seq_name_for_quadruplet.................................................................................................................................................................. 44
-compact........................................................................................................................................................................................................ 44
-clean.............................................................................................................................................................................................................. 44
-maximise..................................................................................................................................................................................................... 44
-do_self.......................................................................................................................................................................................................... 44
-seq_name_for_quadruplet.................................................................................................................................................................. 44
-weight.......................................................................................................................................................................................................... 44
Tree Computation.................................................................................................................................................................................... 45
-distance_matrix_mode......................................................................................................................................................................... 45
-quicktree [CW]......................................................................................................................................................................................... 45
Pair-wise Alignment Computation................................................................................................................................................ 46
-dp_mode..................................................................................................................................................................................................... 46
-ktuple........................................................................................................................................................................................................... 47
-ndiag............................................................................................................................................................................................................. 47
-diag_mode.................................................................................................................................................................................................. 47
-diag_threshold......................................................................................................................................................................................... 47
-sim_matrix.................................................................................................................................................................................................. 47
-matrix [CW]............................................................................................................................................................................................... 48
-nomatch....................................................................................................................................................................................................... 48
-gapopen...................................................................................................................................................................................................... 48
-gapext........................................................................................................................................................................................................... 48
-fgapopen..................................................................................................................................................................................................... 48
-fgapext......................................................................................................................................................................................................... 49
-cosmetic_penalty..................................................................................................................................................................................... 49
-tg_mode....................................................................................................................................................................................................... 49
Weighting Schemes.................................................................................................................................................................................. 49
-seq_weight................................................................................................................................................................................................. 49
Multiple Alignment Computation................................................................................................................................................... 50
-msa_mode................................................................................................................................................................................................... 50
-one2all......................................................................................................................................................................................................... 50
-profile_comparison................................................................................................................................................................................ 50
-profile_mode............................................................................................................................................................................................. 50
Alignment Post-Processing................................................................................................................................................................ 51
-clean_aln..................................................................................................................................................................................................... 51
-clean_threshold....................................................................................................................................................................................... 51
-clean_iteration......................................................................................................................................................................................... 51
-clean_evaluation_mode....................................................................................................................................................................... 51
-iterate........................................................................................................................................................................................................... 51
BLAST Template Selection Parameters...................................................................................... 51
-blast_server................................................................................................................................................................................................ 52
-prot_min_sim............................................................................................................................................................................................ 52
-prot_max_sim............................................................................................................................................................................................ 52
-prot_min_cov............................................................................................................................................................................................. 52
-protein_db.................................................................................................................................................................................................. 52
-pdb_min_sim............................................................................................................................................................................................. 52
-pdb_max_sim............................................................................................................................................................................................ 52
-pdb_min_cov............................................................................................................................................................................................. 53
-pdb_db......................................................................................................................................................................................................... 53
-pdb_type..................................................................................................................................................................................................... 53
CPU Control................................................................................................................................... 53
Multithreading............................................................................................................................................................................................ 53
-multi_core................................................................................................................................................................................................... 53
-n_core........................................................................................................................................................................................................... 54
Limits................................................................................................................................................................................................................ 54
-mem_mode................................................................................................................................................................................................. 54
-ulimit............................................................................................................................................................................................................ 54
-maxlen......................................................................................................................................................................................................... 54
Aligning more than 100 sequences with DPA......................................................................................................................... 54
-maxnseq...................................................................................................................................................................................................... 54
-dpa_master_aln....................................................................................................................................................................................... 55
-dpa_maxnseq............................................................................................................................................................................................ 55
-dpa_min_score1...................................................................................................................................................................................... 55
-dpa_min_score2...................................................................................................................................................................................... 55
-dap_tree [NOT IMPLEMENTED].................................................................................................................................................... 55
Using Structures.......................................................................................................................... 56
Generic............................................................................................................................................................................................................. 56
-mode............................................................................................................................................................................................................. 56
-check_pdb_status.................................................................................................................................................................................... 56
3D Coffee: Using SAP................................................................................................................................................................................ 56
Using/finding PDB templates for the Sequences................................................................................................................... 57
-template_file............................................................................................................................................................................................. 57
-struc_to_use............................................................................................................................................................................................... 59
Multiple Local Alignments........................................................................................................... 59
-domain/-mocca........................................................................................................................................................................................ 59
-start................................................................................................................................................................................................................ 59
-len.................................................................................................................................................................................................................. 60
-scale............................................................................................................................................................................................................... 60
-domain_interactive [Examples]...................................................................................................................................................... 60
Output Control............................................................................................................................. 61
Generic............................................................................................................................................................................................................. 61
Conventions Regarding Filenames................................................................................................................................................. 61
Identifying the Output files automatically.................................................................................................................................. 61
-no_warning............................................................................................................................................................................................... 61
Alignments.................................................................................................................................................................................................... 61
-outfile........................................................................................................................................................................................................... 61
-output........................................................................................................................................................................................................... 61
-outseqweight............................................................................................................................................................................................ 62
-case................................................................................................................................................................................................................ 62
-cpu................................................................................................................................................................................................................. 63
-outseqweight............................................................................................................................................................................................ 63
-outorder [cw]........................................................................................................................................................................................... 63
-inorder [cw].............................................................................................................................................................................................. 63
-seqnos........................................................................................................................................................................................................... 63
Libraries......................................................................................................................................................................................................... 63
-out_lib........................................................................................................................................................................................................... 64
-lib_only........................................................................................................................................................................................................ 64
Trees................................................................................................................................................................................................................. 64
-newtree....................................................................................................................................................................................................... 64
Reliability Estimation................................................................................................................... 64
CORE Computation.................................................................................................................................................................................. 64
-evaluate_mode......................................................................................................................................................................................... 64
Generic Output............................................................................................................................ 65
-run_name................................................................................................................................................................................................... 65
-quiet.............................................................................................................................................................................................................. 65
-align [CW].................................................................................................................................................................................................. 65
APDB, iRMSD and tRMSD Parameters........................................................................................ 65
-quiet [Same as T-Coffee]..................................................................................................................................................................... 66
-run_name [Same as T-Coffee].......................................................................................................................................................... 66
-aln.................................................................................................................................................................................................................. 66
-n_excluded_nb......................................................................................................................................................................................... 66
-maximum_distance................................................................................................................................................................................ 66
-similarity_threshold.............................................................................................................................................................................. 66
-local_mode................................................................................................................................................................................................. 67
-filter............................................................................................................................................................................................................... 67
-print_rapdb [Unsupported].............................................................................................................................................................. 67
-outfile [Same as T-Coffee].................................................................................................................................................................. 67
-color_mode................................................................................................................................................................................................. 67
Environment Variables......................................................................................................................................................................... 68
Output of the .dnd file............................................................................................................................................................................. 69
Permissions.................................................................................................................................................................................................. 69
Other Programs......................................................................................................................................................................................... 69
Parameter files............................................................................................................................ 70
Sequence Name Handling........................................................................................................... 70
Automatic Format Recognition................................................................................................... 71
Structures.................................................................................................................................... 71
RNA Structures............................................................................................................................. 71
Sequences.................................................................................................................................... 71
Alignments................................................................................................................................... 71
Libraries....................................................................................................................................... 72
T-COFFEE_LIB_FORMAT_01............................................................................................................................................................. 72
T-COFFEE_LIB_FORMAT_02............................................................................................................................................................. 73
Library List................................................................................................................................... 73
Substitution matrices.................................................................................................................. 73
ClustalW Style [Deprecated].............................................................................................................................................................. 73
BLAST Format [Recommended]..................................................................................................................................................... 74
Sequences Weights..................................................................................................................... 74
Development............................................................................................................................... 76
profile2list................................................................................................................................................................................................... 76
Command Line List.................................................................................................................................................................................. 76
Please make sure you have agreed with the terms of the license attached to the package before using the T-Coffee package or its documentation. T-Coffee is a freeware open source distributed under a GPL license. This means that there are very little restrictions to its use, either in an academic or a non academic environment.
Our philosophy is that code is meant to be re-used, including ours. No permission is needed for the cut and paste of a few functions, although we are always happy to receive pieces of improved code.
Our philosophy is to insure that as many methods as possible can be used as plug-ins within T-Coffee. Likewise, we will give as much support as possible to anyone wishing to turn T-Coffee into a plug-in for another method. For more details on how to do this, see the plug-in and the plug-out sections of the Tutorial Manual.
Again, you do not need our permission to either use T-Coffee (or your method as a plug-in/out) but if you let us know, we will insure the stability of T-Coffee within your system through future releases.
The current license only allows for the incorporation of T-Coffee in non-commercial pipelines (i.e. where you do not sell the pipeline, or access to it). If your pipeline is commercial, please get in touch with us.
Addresses and Contacts
T-coffee is developed, maintained, monitored, used and debugged by a dedicated team that include or have included:
Cédric Notredame, Paolo di Tomasso, Jean-François Taly, Cedrik Magis, Fabrice Armougom, Des Higgins, Sebastien Moretti, Orla O’Sullivan. Eamon O’Toole, Olivier Poirot, Karsten Suhre, Iain Wallace, Andreas Wilm
We are always very eager to get some user feedback. Please do not hesitate to drop us a line at<EMAIL_ADDRESS>The latest updates of T-Coffee are always available on: www.tcoffee.org . On this address you will also find a link to some of the online T-Coffee servers, including Tcoffee@igs
T-Coffee can be used to automatically check if an updated version is available, however the program will not update automatically, as this can cause endless reproducibility problems.
PROMPT: t_coffee –update
It is important that you cite T-Coffee when you use it. Citing us is (almost) like giving us money: it helps us convincing our institutions that what we do is useful and that they should keep paying our salaries and deliver Donuts to our offices from time to time (Not that they ever did it, but it would be nice anyway).
Cite the server if you used it, otherwise, cite the original paper from 2000 (No, it was never named "T-Coffee 2000").
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Related Articles, Links
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T-Coffee: A novel method for fast and accurate
multiple sequence alignment. |
Other useful publications include:
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Related Articles, Links
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CaspR: a web server for automated molecular
replacement using homology modelling. |
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Related Articles, Links
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3DCoffee@igs: a web server for combining sequences
and structures into a multiple sequence alignment. |
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Related Articles, Links
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3DCoffee: combining protein sequences and
structures within multiple sequence alignments. |
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Related Articles, Links
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Tcoffee@igs: A web server for computing,
evaluating and combining multiple sequence alignments. |
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Related Articles, Links
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Mocca: semi-automatic method for domain hunting. |
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Related Articles, Links
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T-Coffee: A novel method for fast and accurate
multiple sequence alignment. |
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Related Articles, Links
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COFFEE: an objective function for multiple
sequence alignments. |
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Related Articles, Links |
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Mocca: semi-automatic method for domain
hunting. |
http://www.tcoffee.org/Publications/Pdf/core.pp.pdf
We do not mean to steal code, but we will always try to re-use pre-existing code whenever that code exists, free of copyright, just like we expect people to do with our code. However, whenever this happens, we make a point at properly citing the source of the original contribution. If ever you recognize a piece of your code improperly cited, please drop us a note and we will be happy to correct that.
In the mean time, here are some important pieces of code from other packages that have been incorporated within the T-Coffee package. These include:
-The Sim algorithm of Huang and Miller that given two sequences computes the N best scoring local alignments.
-The tree reading/computing routines are taken from the ClustalW Package, courtesy of Julie Thompson, Des Higgins and Toby Gibson (Thompson, Higgins, Gibson, 1994, 4673-4680,vol. 22, Nucleic Acid Research).
-The implementation of the algorithm for aligning two sequences in linear space was adapted from Myers and Miller, in CABIOS, 1988, 11-17, vol. 1)
-Various techniques and algorithms have been implemented. Whenever relevant, the source of the code/algorithm/idea is indicated in the corresponding function.
-64 Bits compliance was implemented by Benjamin Sohn, Performance Computing Center Stuttgart (HLRS), Germany
-David Mathog (Caltech) provided many fixes and useful feedback for improving the code and making the whole soft behaving more rationally
-Prof David Jones (UCL) reported and corrected the PDB1K bug (now t_coffee/sap can align PDB sequences longer than 1000 AA).
-Johan Leckner reported several bugs related to the treatment of PDB structures, insuring a consistent behavior between version 1.37 and current ones.
Installation of The T-Coffee Packages
T-Coffee is a complex package that interacts with many other third part software. If you only want a standalone version of T-Coffee, you may install that package on its own. If you want to use a most sophisticated flavor (3dcoffee, expresso, rcofeee, etc...), the installer will try to install all the third party packages required.
Note that since version 7.56, T-Coffee will use 'on demand' installation and install the third party packages it needs *when* it needs them. This only works for packages not requiring specific licenses and that can be installed by the regular installer. Please let us know if you would like another third party package to be included.
Whenver on-demand installation or automated installation fails because of unforessen system specificities, users should install the third party package manually. This documentation gives some tips we have found useful, but users are encouraged to check the original documentation.
We now recommend that you use the automated installer provided for UNIX or Linux platforms.
Click on tge .dmg file
rpm –i <rmp>
You need to have: gcc, g77, CPAN and an internet connection and your root password. You will also need the two following perl modules: LWP and XML::Simple. These are needed if you want to use the web services provided by the EBI via REST (http://www.ebi.ac.uk/Tools/webservices/tutorials/02_rest)
1. gunzip t_coffee.tar.gz
2. tar -xvf t_coffee.tar
3. cd t_coffee
4. ./install t_coffee
This installation will only install the stand alone T-Coffee. If you want to install a specific mode of T-Coffee, you may try the following commands that will try to gather all the necessary third party packages. Note that a package already found on your system will not be re-installed.
./install t_coffee
./install mcoffee
./install 3dcoffee
./install rcoffee
./install psicoffee
Or even
./install all
-All the corresponding executables will be downloaded automatically and installed in
$HOME/.t_coffee/plugins
-if you executables are in a different location, give it to T-Coffee using the -plugins flag.
-If the installation of any of the companion package fails, you should install it yourself using the provided link (see below) and following the authors instructions.
-If you have not managed to install SOAP::Lite, you can re-install it later (from anywhere) following steps 1-2.
-This procedure attempts 3 things: installing and Compiling T-Coffee (C program), Installing and compiling TMalign (Fortran), Installing and compiling XML::Simple
-If you have never installed a perl module before, CPAN will ask you many questions: say Yes to all
-If everything went well, the procedure has created in the bin directory two executables: t_coffee and TMalign (Make sure these executables are on your $PATH!)
Install Cygwin
Download The Installer (NOT Cygwin/X)
Click on view to list ALL the packages
Select: gcc-core, make, wget
Optional: ssh, xemacs, nano
Run mkpasswd in Cywin (as requested when you start cygwin)
Install T-Coffee within Cygwin using the Unix procedure
Make sure you have the Developer's kit installed (compilers and makefile)
Follow the Unix Procedure
In order to run, T-Coffee must have a value for the http_proxy and for the E-mail. In order to do so you can either:
export the following values:
export http_proxy_4_TCOFFEE="proxy" or "" if no proxy
export EMAIL_4_TCOFFEE="your email"
OR
modify the file ~/.t_coffee/t_coffee_env
OR
add to your command line: t_coffee …. -proxy=<proxy> -email=<email
if you have no proxy: t_coffee … -proxy -email=<email>
Assuming you have a standard PDB installation in your file system
setenv (or export) PDB_DIR <abs path>/data/structures/all/pdb/
OR
setenv (or export) PDB_DIR <abs path>/structures/divided/pdb/
If you do not have PDB installed, don't worry, t_coffee will go and fetch any structure it needs directly from the PDB repository. It will simply be a bit slower than if you had PDB locally.
BLAST is a program that search sequence databases for homologues of a query sequence. It works for proteins and Nucleic Acids. In theory BLAST is just a package like any, but in practice things are a bit more complex. To run well, BLST requires up to date databases (that can be fairly large, like NR or UNIPROT) and a powerful computer.
Fortunately, an increasing number of institutes or companies are now providing BLAST clients that run over the net. It means that all you need is a small program that send your query to the big server and gets the results back. This prevents you from the hassle of installing and maintaining BLAST, but of course it is less private and you rely on the network and the current load of these busy servers.
Thanks to its interaction with BLAST, T-Coffee can gather structures and protein profiles and deliver an alignment significantly more accurate than the default you would get with T-Coffee or any similar method.
Let us go through the various modes available for T-Coffee
The most accurate modes of T-Coffe scan the databases for templates that they use to align the sequences. There are currently two types of templates for proteins:
structures (PDB) that can be found by a blastp against the PDB database and profiles that can be constructed with eiether a blastp or a psiblast against nr or uniprot.
These templates are automatically built if you use:
t_coffee <yourseq> -mode expresso
that fetches aand uses pdb templates, or
t_coffee <your seq> -mode psicoffee
that fetches and uses profile templates, or
t_coffee <your seq> -mode accurate
that does everything and tries to use the best template. Now that you see why it is useful let's see how to get BLAST up and running, from the easy solution to tailor made ones.
This is by far the easiest (and the default mode). The perl clients are already incorporated in T-Coffee and all you need is are the proper perl library. In theory, T-Coffee should have already installed these libraries during the standard installation. Yet, this requires having toot access. It really is worth the effort, since the EBI is providing one of the best webservice available around, and most notably, the only public psiblast via a web service.
Whenever you use a T-Coffee mode requiring Blast access, it will ask you for an authentification E-mail. Be Careful! If you provide a fake E-mail, the EBI may suspend the service for all machines associated with your IP address (that could mean your entire lab, or entire institute, or even the entire country or, but I doubt it, the whole universe).
The NCBI is the next best alternative. In my hand it was always a bit slower and most of all, it does not incorporate PSI-BLAST (as a web sevice). A big miss. The NCBI web blast client is a small executable that you should install on your system following the instructions given on this link
ftp://ftp.ncbi.nih.gov/blast/executables/LATEST
Simply go for netbl, download the executable that corresponds to your architecture (cygwin users should go for the win executable). Despite all the files that come along the executable blastcl3 is a stand alone executable that you can safely move to your $BIN.
All you will then need to do is to make sure that T-Coffee uses the right client, when you run it.
-blast_server=NCBI
No need for any E-mail here, but you don't get psiblast, and whenever T-Coffee wants to use it, blastp will be used instead.
You may have your own client (lucky you). If that is so, all you need is to make sure that this client is complient with the blast command line. If your client is named foo.pl, all you need to to is run T-Coffee with
-blast_server=CLIENT_foo.pl
Foo will be called as if it were blastpgp, and it is your responsability to make sure it can handle the following command line:
foo.pl -p <method> -d <db> -i <infile> -o <outfile> -m 7
method can either be blastp or psiblast.
infile is a FASTA file
-m7 triggers the XML output. T-Coffee is able to parse both the EBI XML output and the NCBI XML output.
If foo.pl behaves differently, the easiest will probably be to write a wrapper around it so that wrapped_foo.pl behaves like blastpgp
If you have blastpgp installed, you can run it instead of the remote clients by using:
-blast_server=LOCAL
The documentation for blastpgp can be found on:
www.ncbi.nlm.nih.gov/staff/tao/URLAPI/blastpgp.html
and the package is part of the standard BLAST distribution
ftp://ftp.ncbi.nih.gov/blast/executables/LATEST
Depending on your system, your own skills, your requirements and on more parameters than I have fingers to count, installing a BLAST server suited for your needs can range from a 10 minutes job to an achivement spread over several generations. So at this point, you should roam the NCBI website for suitable information.
If you want to have your own BLAST server to run your own databases, you should know that it is possible to control both the database and the program used by BLAST:
-protein_db: will specify the database used by all the psi-blast modes
-pdb_db: will specify the database used by the pdb modes
Note that T-Coffee is compliant with BLAST+, the latest NCBI Blast.
Blast+ is tghe latest NCBI Blast. IT is easier to install. A default installation should be compliant with a default T-Coffee installation.
For those of you using cygwin, be careful. While cygwin behaves like a UNIX system, the BLAST executable required for cygwin (win32) is expecting WINDOWS path and not UNIX path. This has three important consequences:
1- the ncbi file declaring the Data directory must be:
C:WINDOWS//ncbi.init [at the root of your WINDOWS]
2- the address mentionned with this file must be WINDOWS formated, for instance, on my system:
Data=C:\cygwin\home\notredame\blast\data
3- When you pass database addresses to BLAST, these must be in Windows format:
-protein_db="c:/somewhere/somewhereelse/database"
(using the slash (/) or the andtislash (\) does not matter on new systems but I would reommand against incorporating white spaces.
T-Coffee is meant to interact with as many packages as possible, either for aligning or using predictions. If you type
t_coffee
You will receive a list of supported packages that looks like the next
table. In theory, most of these packages can be installed by T-Coffee and we welcome any reasonnable request.
 If you do not have any structural information, it is by far the most accurate mode of T-Coffee. To use it, you must have SOAP installed so that the EBI BLAST client can run on your system.
It is a bit slow, but really worth it if your sequences are hard to align and if the accuracy of your alignment is important.
To use this mode, try:
t_coffee <yoursequence> -mode psicoffee
Note that because PSI-BLAST is time consuming, T-Coffee stores the runs in its cache (./tcoffee/cache) so that it does not need to be re-run. It means that if you re-align your sequences (or add a few extra sequences), things will be considerably faster.
If your installation procedure has managed to compile TMalign, and if T-Coffee has access to the EBI BLAST server (or any other server) you can also do the following:
t_coffee <yoursequence> -mode expresso
That will look for structural templates. And if both these modes are running fine, then you are ready for the best, the "crème de la crème":
t_coffee <yoursequence> -mode accurate
M-Coffee is a special mode of T-Coffee that makes it possible to combine
the output of many multiple sequence alignment packages.
In the T-Coffee distribution, type:
./install mcoffee
In theory, this command should download and install every required package. If, however, it fails, you should switch to the manual installation (see next).
By default these packages will be in
$HOME/.t_coffee/plugins
If you want to have these companion packages in a different directory, you can either set the environement variable
setenv PLUGINS_4_TCOFFEE=<plugins dir>
Or use the command line flag -plugin (over-rides every other setting)
t_coffee ... -plugins=<plugins dir>
M-Coffee requires a standard T-Coffee installation (c.f. previous section) and the following packages to be installed on your system:
Package Where From
==========================================================
ClustalW can interact with t_coffee
----------------------------------------------------------
Poa http://www.bioinformatics.ucla.edu/poa/
----------------------------------------------------------
Muscle http://www.drive5.com
----------------------------------------------------------
ProbCons http://probcons.stanford.edu/
ProbConsRNA http://probcons.stanford.edu/
----------------------------------------------------------
MAFFT http://www.biophys.kyoto-u.ac.jp/~katoh/programs/align/mafft/
----------------------------------------------------------
Dialign-T http://dialign-t.gobics.de/
Dialign-TX http://dialign-tx.gobics.de/
----------------------------------------------------------
PCMA ftp://iole.swmed.edu/pub/PCMA/
----------------------------------------------------------
kalign http://msa.cgb.ki.se
----------------------------------------------------------
amap http://bio.math.berkeley.edu/amap/
-----------------------------------------------------------
proda_msa http://bio.math.berkeley.edu/proda/
-----------------------------------------------------------
prank_msa http://www.ebi.ac.uk/goldman-srv/prank/
In our hands all these packages where very straightforward to compile and install on a standard cygwin or Linux configuration. Just make sure you have gcc, the C compiler, properly installed.
Once the package is compiled and ready to use, make sure that the executable is on your path, so that t_coffee can find it automatically. Our favorite procedure is to create a bin directory in the home. If you do so, make sure this bin is in your path and fill it with all your executables (this is a standard Unix practice).
If for some reason, you do not want this directory to be on your path, or you want to specify a precise directory containing the executables, you can use:
export PLUGINS_4_TCOFFEE=<dir>
By default this directory is set to $HOME/.t_coffee/plugins/$OS, but you can over-ride it with the environement variable or using the flag:
t_coffee ...-plugins=<dir>
If you cannot, or do not want to use a single bin directory, you can set the following environment variables to the absolute path values of the executable you want to use. Whenever they are set, these variables will supersede any other declaration. This is a convenient way to experiment with multiple package versions.
POA_4_TCOOFFEE
CLUSTALW_4_TCOFFEE
POA_4_TCOFFEE
TCOFFEE_4_TCOFFEE
MAFFT_4_TCOFFEE
MUSCLE_4_TCOFFEE
DIALIGNT_4_TCOFFEE
PRANK_4_TCOFFEE
DIALIGNTX_4_TCOFFEE
For three of these packages, you will need to copy some of the files in a special T-Coffee directory.
cp POA_DIR/* ~/.t_coffee/mcoffee/
cp DIALIGN-T/conf/* ~/.t_coffee/mcoffee
cp DIALIGN-TX/conf/* ~/.t_coffee/mcoffee
Note that the following files are enough for default usage:
BLOSUM.diag_prob_t10 BLOSUM75.scr blosum80_trunc.mat
dna_diag_prob_100_exp_330000 dna_diag_prob_200_exp_110000
BLOSUM.scr BLOSUM90.scr dna_diag_prob_100_exp_110000
dna_diag_prob_100_exp_550000 dna_diag_prob_250_exp_110000
BLOSUM75.diag_prob_t2 blosum80.mat dna_diag_prob_100_exp_220000
dna_diag_prob_150_exp_110000 dna_matrix.scr
If you would rather have the mcoffee directory in some other location, set the MCOFFEE_4_TCOFFEE environement variable to the propoer directory:
setenv MCOFFEE_4_TCOFFEE <directory containing mcoffee files>
APDB and iRMSD are incorporated in T-Coffee. Once t_coffee is installed, you can invoque these programs by typing:
t_coffee
–other_pg apdb
t_coffee –other_pg
irmsd
tRMSD comes along with t_coffee but it also requires the package phylip in order to be functional. Phylip can be obtained from:
Package Function
===================================================
---------------------------------------------------
Phylip Phylogenetic tree computation
evolution.genetics.washington.edu/phylip.html
---------------------------------------------------
t_coffee –other_pg trmsd
Seq_reformat is a reformatting package that is part of t_coffee. To use it (and see the available options), type:
t_coffee –other_pg seq_reformat
Extract_from_pdb is a PDB reformatting package that is part of t_coffee. To use it (and see the available options), type.
t_coffee –other_pg extract_from_pdb –h
Extract_from_pdb requires wget in order to automatically fetch PDB structures.
3D-Coffee/Expresso is a special mode of
T-Coffee that makes it possible to combine sequences and structures. The main
difference between Expresso and 3D-Coffee is that Expresso fetches the
structures itself.
In the T-Coffee distribution, type:
./install expresso
OR
./install 3dcoffee
In theory, this command should download and install every required package (except fugue). If, however, it fails, you should switch to the manual installation (see next).
In order to make the most out of T-Coffee, you will need to install the following packages (make sure the executable is named as indicated below):
Package Function
===================================================
---------------------------------------------------
wget 3DCoffee
Automatic Downloading of Structures
---------------------------------------------------
sap structure/structure comparisons
(obtain it from W. Taylor, NIMR-MRC).
---------------------------------------------------
TMalign zhang.bioinformatics.ku.edu/TM-align/
---------------------------------------------------
mustang www.cs.mu.oz.au/~arun/mustang/
---------------------------------------------------
wublastclient www.ebi.ac.uk/Tools/webservices/clients/wublast
---------------------------------------------------
Blast www.ncbi.nih.nlm.gov
---------------------------------------------------
Fugue* protein to structure alignment program
http://www-cryst.bioc.cam.ac.uk/fugue/download.html
***NOT COMPULSORY***
Once the package is installed, make sure make sure that the executable is on your path, so that t_coffee can find it automatically.
The wublast client makes it possible to run BLAST at the EBI without having to install any database locally. It is an ideal solution if you are only using expresso occasionally.
Uses a standard fugue installation. You only need to install the following packages:
joy, melody, fugueali, sstruc, hbond
If you have root privileges, you can install the common data in:
cp fugue/classdef.dat /data/fugue/SUBST/classdef.dat
otherwise
Setenv MELODY_CLASSDEF=<location>
Setenv MELODY_SUBST=fugue/allmat.dat
All the other configuration files must be in the right location.
R-Coffee is a special mode able to align RNA
sequences while taking into account their secondary structure.
In the T-Coffee distribution, type:
./install rcoffee
In theory, this command should download and install every required package (except consan). If, however, it fails, you should switch to the manual installation (see next).
R-Coffee only requires the package Vienna to be installed, in order to compute multiple sequence alignments. To make the best out of it, you should also have all the packages required by M-Coffee
Package Function
===================================================
---------------------------------------------------
consan R-Coffee
Computes highly accurate pairwise Alignments
***NOT COMPULSORY***
selab.janelia.org/software/consan/
---------------------------------------------------
RNAplfold Computes RNA secondary Structures
www.tbi.univie.ac.at/~ivo/RNA/
---------------------------------------------------
probconsRNA probcons.stanford.edu/
---------------------------------------------------
M-Coffee T-Coffee and the most common MSA Packages
(cf M-Coffee in this installation guide)
Follow the installation procedure, but make sure you rename the probcons executable into probconsRNA.
In order to insure a proper interface beween consan and R-Coffee, you must make sure that the file mix80.mod is in the directory ~/.t_coffee/mcoffee or in the mcoffee directory otherwise declared.
Quick Start
We only give you the very basics here. Please use the Tutorial for more detailed information on how to use our tools.
IMPORTANT: All the files mentionned here (sampe_seq...) can be found in the example directory of the distribution.
Write your sequences in the same file (Swiss-prot, Fasta or Pir) and type.
PROMPT: t_coffee sample_seq1.fasta
This will output two files:
sample_seq1.aln: your Multiple Sequence Alignment
sample_seq1.dnd: The Guide tree (newick Format)
IMPORTANT:
In theory nucleic acids should be automatically detected and the default methods should be adapted appropriately. However, sometimes this may fail, either because the sequences are too short or contain too many ambiguity codes.
When this happens, you are advised to explicitly set the type of your sequences
NOTE: the –mode=dna is not needed or supported anymore
PROMPT: t_coffee sample_dnaseq1.fasta –type=dna
M-Coffee is a Meta version of T-Coffee that makes it possible to combine the output of at least eight packages (Muscle, probcons, poa, dialignT, mafft, clustalw, PCMA and T-Coffee).
If all these packages are already installed on your machine. You must:
1-set the following environment variables
export POA_DIR=[absolute path of the POA installation dir]
export DIALIGNT_DIR=[Absolute path of the DIALIGN-T/conf
Once this is done, write your sequences in a file and run: same file (Swiss-prot, Fasta or Pir) and type.
PROMPT: t_coffee sample_seq1.fasta –mode mcoffee
If the program starts complaining one package or the other is missing, this means you will have to go the hard way and install all these packages yourself... Proceed to the M-Coffee section for more detailed instructions.
If you have installed the EBI wublast.pl client, Expresso will BLAST your sequences against PDB, identify the best targets and use these to align your proteins.
PROMPT: t_coffee sample_seq1.fasta –mode expresso
If you did not manage to install all the required structural packages for Expresso, like Fugue or Sap, you can still run expresso by selecting yourself the structural packages you want to use. For instance, if you'd rather use TM-Align than sap, try:
PROMPT: t_coffee sample_seq1.fasta –template_file EXPRESSO -method TMalign_pair
R-Coffee can be used to align RNA sequences, using their RNApfold predicted secondary structures. The best results are obtained by using the consan pairwise method. If you have consan installed:
t_coffee sample_rnaseq1.fasta –special_mode rcoffee_consan
This will only work if your sequences are short enough (less than 200 nucleotides). A good alternative is the rmcoffee mode that will run Muscle, Probcons4RNA and MAfft and then use the secondary structures predicted by RNApfold.
PROMPT: t_coffee sample_rnaseq1.fasta –mode mrcoffee
If you want to decide yourself which methods should be combined by R-Coffee, run:
PROMPT: t_coffee sample_rnaseq1.fasta –mode rcoffee -method lalign_id_pair slow_pair
All you need is a file containing the alignment of sequences with a known structure. These sequences must be named according to their PDB ID, followed by the chain index ( 1aabA for instance). All the sequences do not need to have a known structure, but at least two need to have it.
Given the alignment:
PROMPT: t_coffee –other_pg irmsd -aln 3d_sample4.aln
tRMSD is a structure based clustering method using the iRMSD to drive the clustering. The T-RMSD supports all the parameters supported by iRMSD or APDB.
PROMPT: t_coffee –other_pg trmsd -aln 3d_sample5.aln -template_file 3d_sample5.template_list
3d_sample5.aln is a multiple alignment in which each sequence has a known structure. The file 3d_sample5.template_list is a fasta like file declaring the structure associated with each sequence, in the form:
> <seq_name> _P_ <PDB structure file or name>
******* 3d_sample5.template_list ********
>2UWI-3A _P_ 2UWI-3.pdb
>2UWI-2A _P_ 2UWI-2.pdb
>2UWI-1A _P_ 2UWI-1.pdb
>2HEY-4R _P_ 2HEY-4.pdb
...
**************************************
The program then outputs a series of files
Template Type: [3d_sample5.template_list] Mode Or File:
[3d_sample5.template_list] [Start]
[Sample
Columns][TOT= 51][100
%][ELAPSED TIME: 0
sec.]
[Tree
Cmp][TOT= 13][ 92 %][ELAPSED
TIME: 0 sec.]
#### File
Type= TreeList Format= newick Name=
3d_sample5.tot_pos_list
#### File
Type=
Tree Format=
newick Name= 3d_sample5.struc_tree10
#### File
Type=
Tree Format=
newick Name= 3d_sample5.struc_tree50
#### File
Type=
Tree Format=
newick Name= 3d_sample5.struc_tree100
#### File
Type= Colored MSA Format= score_html Name= 3d_sample5.struc_tree.html
3d_sample5.tot_pos_list is a list of the tRMSD tree associated with every position.
3d_sample5.struc_tree100 is
a consensus tree (phylip/consense) of the trees contained in the previous file.
This file is the default output
3d_sample5.struc_tree10 is a consensus tree (phylip/consense) of the 10% trees having the higest average agreement with the rest
3d_sample5.struc_tree10 is a consensus tree (phylip/consense) of the 50% trees having the higest average agreement with the rest
3d_sample5.html is a colored version of the output showing in red the positions that give the highest support to 3d_sample5.struc_tree100
Write your sequences in the same file (Swiss-prot, Fasta or Pir) and type.
PROMPT: t_coffee –other_pg mocca sample_seq1.fasta
This command output one files (<your sequences>.mocca_lib) and starts an interactive menu.
Recent Modifications
Warning: This log of recent modifications is not as thorough and accurate as it should be.
-9.86 New data structure for the primary library that results in highly improved running times for mcoffee and significantly decreased memory usage.
-5.80 Novel assembly algorithm (linked_pair_wise) and the primary library is now made of probcons style pairwise alignments (proba_pair)
-4.30 and upward: the FAQ has moved into a new tutorial document
-4.30 and upward: -in has will be deprecated and replaced by the flags: -profile,-method,-aln,-seq,-pdb
-4.02: -mode=dna is still available but not any more needed or supported. Use type=protein or dna if you need to force things
-3.28: corrected a bug that prevents short sequences from being correctly aligned
-Use of @ as a separator when specifying methods parameters
-The most notable modifications have to do with the structure of the input. From version 2.20, all files must be tagged to indicate their nature (A: alignment, S: Sequence, L: Library…). We are becoming stricter, but that’s for your own good…
Another important modification has to do with the flag -matrix: it now controls the matrix being used for the computation
Reference Manual
This reference manual gives a list of all the flags that can be used to modify the behavior of T-Coffee. For your convenience, we have grouped them according to their nature. To display a list of all the flags used in the version of T-Coffee you are using (along with their default value), type:
PROMPT: t_coffee
Or
PROMPT: t_coffee –help
Or
PROMPT: t_coffee
–help –in
Or any other parameter
It is possible to modify T-Coffee’s behavior by setting any of the following environment variables. On the bash shell, use export VAR=”value”. On the cshell, use set $VAR=”xxx”
Sets the http_proxy and HTTP_proxy values used by T-Coffee.
These values get supersede http_proxy and HTTP_proxy. http_proxy_4_TCOFFEE gets superseded by the command line values (-proxy and -email)
If you have no proxy, just set this value to an empty string.
Set the E-mail values provided to web services called upon by T-Coffee.
Can be over-riden by the flag -email.
By default this variable is set to $HOME/.t_coffee. This is where T-Coffee expects to find its cache, tmp dir and possibly any temporary data stored by the program.
By default this variable is set to $HOME/.t_coffee/tmp. This is where T-Coffee stores temporary files.
By default this variable is set to $HOME/.t_coffee/cache. This is where T-Coffee stores any data expensive to obtain: pdb files, sap alignments....
By default all the companion packages are searched in the directory DIR_4_TCOFFEE/plugins/<OS>. This variable overrides the default. This variable can also be overriden by the -plugins T-Coffee flag
By default this variable is no set. Set it if you do not want the program to generate a verbose error output file (useful for running a server).
Indicate the location of your local PDB installation.
Suppresses all the warnings.
Sets:
DIR_4_TCOFFEE
CACHE_4_TCOFFEE
TMP_4_TCOFFEE
PLUGINS_4_TCOFFEE
To the same unique value. The string MUST be a valid directory
T-Coffee can have its own environment file. This environment is kept in a file named $HOME/.t_coffee/t_coffee_env and can be edited. The value of any legal variable can be modified through that file. For instance, here is an example of a configuration file when not requiring a proxy.
http_proxy_4_TCOFFEE=
<EMAIL_ADDRESS>If you want to use a specific configuration file:
t_coffee … -setenv ENV_4_TCOFFEE=<location>
In general, you can set any environment variable using the –setenv flag. You can also simply do an export:
export ENV_4_TCOFFEE=<location>
IMPORTANT:
export > –setenv > -proxy, -email > t_coffee_env > default environment
Note that when you use –setenv for PATH, the value you provide is concatenated TO THE BEGINNING of the current PATH value. This way you can force T-Coffee to use a specific version of an aligner.
You can use any kind of separator you want (i.e. ,; <space>=). The syntax used in this document is meant to be consistent with that of ClustalW. However, in order to take advantage of the automatic filename compleation provided by many shells, you can replace “=” and “,” with a space.
T-Coffee is not POSIX compliant (sorry L).
There are many ways to enter parameters in T-Coffee, see the -parameter flag in
Parameters Priority
In general you will not need to use these complicated parameters. Yet, if you find yourself typing long command lines on a regular basis, it may be worth reading this section.
One may easily feel confused with the various manners in which the parameters can be passed to t_coffee. The reason for these many mechanisms is that they allow several levels of intervention. For instance, you may install t_coffee for all the users and decide that the defaults we provide are not the proper ones… In this case, you will need to make your own t_coffee_default file.
Later on, a user may find that he/she needs to keep re-using a specific set of parameters, different from those in t_coffee_default, hence the possibility to write an extra parameter file: parameters. In summary:
-parameters > prompt parameters > -t_coffee_defaults > -mode
This means that -parameters supersede all the others, while parameters provided via -special mode are the weakest.
If no flag is used <your sequence> must be the first argument. See format for further information.
PROMPT: t_coffee sample_seq1.fasta
Which is equivalent to
PROMPT: t_coffee Ssample_seq1.fasta
When you do so, sample_seq1 is used as a name prefix for every file the program outputs.
Usage: -parameters=parameters_file
Default: no parameters file
Indicates a file containing extra parameters. Parameters read this way behave as if they had been added on the right end of the command line that they either supersede(one value parameter) or complete (list of values). For instance, the following file (parameter.file) could be used
*******sample_param_file.param********
-in=Ssample_seq1.fasta,Mfast_pair
-output=msf_aln
**************************************
Note: This is one of the
exceptions (with –infile) where the identifier tag (S,A,L,M…) can be
omitted. Any dataset provided this way will be assumed to be a sequence (S).
These exceptions have been designed to keep the program compatible with
ClustalW.
Note: This parameter file
can ONLY contain valid parameters. Comments are not allowed. Parameters passed
this way will be checked like normal parameters.
Used with:
PROMPT: t_coffee -parameters=sample_param_file.param
Will cause t_coffee to apply the fast_pair method onto to the sequences contained in sample_seq.fasta. If you wish, you can also pipe these arguments into t_coffee, by naming the parameter file "stdin" (as a rule, any file named stdin is expected to receive its content via the stdin)
cat sample_param_file.param | t_coffee -parameters=stdin
Usage: -t_coffee_defaults=<file_name>
Default: not used.
This flag tells the program to use some default parameter file for t_coffee. The format of that file is the same as the one used with -parameters. The file used is either:
1. <file name> if a name has been specified
2. ~/.t_coffee_defaults if no file was specified
3. The file indicated by the environment variable TCOFFEE_DEFAULTS
Usage: -mode= hard coded mode
Default: not used.
It indicates that t_coffee will use some hard coded parameters. These include:
quickaln: very fast approximate alignment
dali: a mode used to combine dali pairwise alignments
evaluate: defaults for evaluating an alignment
3dcoffee: runs t_coffee with the 3dcoffee parameterization
Other modes exist that are not yet fully supported
Usage: -score
Default: not used
Toggles on the evaluate mode and causes t_coffee to evaluates a precomputed alignment provided via -infile=<alignment>. The flag -output must be set to an appropriate format (i.e. -output=score_ascii, score_html or score_pdf). A better default parameterization is obtained when using the flag -mode=evaluate.
Usage: -evaluate
Default: not used
Replaces –score. This flag toggles on the evaluate mode and causes t_coffee to evaluates a pre-computed alignment provided via -infile=<alignment>. The flag -output must be set to an appropriate format (i.e. -output=score_ascii, score_html or score_pdf).
The main purpose of –evaluate is to let you control every aspect of the evaluation. Yet it is advisable to use pre-defined parameterization: mode=evaluate.
PROMPT: t_coffee –infile=sample_aln1.aln -mode=evaluate
PROMPT: t_coffee –infile=sample_seq1.aln –in Lsample_lib1.tc_lib –mode=evaluate
Usage: -convert
Default: turned off
Toggles on the conversion mode and causes T-Coffee to convert the sequences, alignments, libraries or structures provided via the -infile and -in flags. The output format must be set via the -output flag. This flag can also be used if you simply want to compute a library (i.e. you have an alignment and you want to turn it into a library).
This flag is ClustalW compliant.
Usage: -do_align
Default: turned on
Usage: -version
Default: not used
Returns the current version number
Usage: -proxy=<proxy>
Default: not used
Sets the proxy used by HTTP_proxy AND http_proxy. Setting with the propmpt supersedes ANY other setting.
Note that if you use no proxy, you should set
-proxy
Usage: -email=<email>
Default: not used
Sets your email value as provided to web services
Usage: -check_configuration
Default: not used
Checks your system to determine whether all the programs T-Coffee can interact with are installed.
Usage: -cache=<use, update, ignore, <filename>>
Default: -cache=use
By default, t_coffee stores in a cache directory, the results of computationally expensive (structural alignment) or network intensive (BLAST search) operations.
Usage: -update
Default: turned off
Causes a wget access that checks whether the t_coffee version you are using needs updating.
Usage: -full_log=<filename>
Default: turned off
Causes t_coffee to output a full log file that contains all the input/output files.
Usage: -plugins=<dir>
Default: default
Specifies the directory in which the companion packages (other multiple aligners used by M-Coffee, structural aligners, etc…) are kept as an alternative, you can also set the environment variable PLUGINS_4_TCOFFEE
The default is ~/.t_coffee/plugins/
Usage: -other_pg=<filename>
Default: turned off
Some rumours claim that Tetris is embedded within T-Coffee and could be ran using some special set of commands. We wish to deny these rumours, although we may admit that several interesting reformatting programs are now embedded in t_coffee and can be ran through the –other_pg flag.
PROMPT: t_coffee –other_pg=seq_reformat
PROMPT: t_coffee –other_pg=unpack_all
PROMPT: t_coffee –other_pg=unpack_extract_from_pdb
To remain compatible with ClustalW, it is possible to indicate the sequences with this flag
PROMPT: t_coffee -infile=sample_seq1.fasta
Note: Common multiple sequence
alignments format constitute a valid input format.
Note: T-Coffee
automatically removes the gaps before doing the alignment. This behaviour is different from that of ClustalW
where the gaps are kept.
-in (Cf –in from the Method and Library Input section)
Usage: -get_type
Default: turned off
Forces t_coffee to identify the sequences type (PROTEIN, DNA).
Usage: -type=DNA ¦ PROTEIN¦ DNA_PROTEIN
Default: -type=<automatically set>
This flag sets the type of the sequences. If omitted, the type is guessed automatically. This flag is compatible with ClustalW.
Usage: -seq=[<P,S><name>,]
Default: none
-seq is now the recommended flag to provide your sequences. It behaves mostly like the -in flag.
Usage: -seq_source=<ANY or _LS or LS >
Default: ANY.
You may not want to combine all the provided sequences into a single sequence list. You can do by specifying that you do not want to treat all the –in files as potential sequence sources.
-seq_source=_LA indicates that neither sequences provided via the A (Alignment) flag or via the L (Library flag) should be added to the sequence list.
-seq_source=S means that only sequences provided via the S tag will be considered. All the other sequences will be ignored.
Note: This flag is mostly designed for
interactions between T-Coffee and T-CoffeeDPA (the large scale version of
T-Coffee).
Usage: -pdb=<pdbid1>,<pdbid2>…[Max 200]
Default: None
Reads or fetch a pdb file. It is possible to specify a chain or even a sub-chain:
PDBID(PDB_CHAIN)[opt] (FIRST,LAST)[opt]
It is also possible to input structures via the –in flag. In that case, you will need to use the TAG identifier:
-in Ppdb1 Ppdb2…
Usage: -usetree=<tree file>
Default: No file specified
Format: newick tree format (ClustalW Style)
This flag indicates that rather than computing a new dendrogram, t_coffee must use a pre-computed one. The tree files are in phylips format and compatible with ClustalW. In most cases, using a pre-computed tree will halve the computation time required by t_coffee. It is also possible to use trees output by ClustalW, Phylips and any other program.
The -in Flag and its
Identifier TAGS
<-in> is the real grinder of T-Coffee. Sequences, methods and alignments all pass through so that T-Coffee can turn it all into a single list of constraints (the library). Everything is done automatically with T-Coffee going through each file to extract the sequences it contains. The methods are then applied to the sequences. Pre-compiled constraint list can also be provided. Each file provided via this flag must be preceded with a symbol (Identifier TAG) that indicates its nature to T-Coffee. The TAGs currently supported are the following:
P PDB structure
S for sequences (use it as well to treat an MSA as unaligned sequences)
M Methods used to build the library
L Pre-computed T-Coffee library
A Multiple Alignments that must be turned into a Library
X Substitution matrices.
R Profiles. This is a legal multiple alignments that will be treated as single sequences (the sequences it contains will not be realigned).
If you do not want to use the TAGS, you will need to use the following flags in replacement of -in. Do not use the TAGS when using these flags:
-aln Alignments (A)
-profile Profiles (R)
-method Method (M)
-seq Sequences (S)
Usage: -in=[<P,S,A,L,M,X><name>,]
Default: -in=Mlalign_id_pair,Mclustalw_pair
Note: -in can be replaced with the combined usage of -aln, iprofile, .pdb, .lib, -method.
See the box for an explanation of the -in flag. The following argument passed via -in
PROMPT: t_coffee -in=Ssample_seq1.fasta,Asample_aln1.aln,Asample_aln2.msf,Mlalign_id_pair,Lsample_lib1.tc_lib –outfile=outaln
This command will trigger the following chain of events:
1-Gather all the sequences
Sequences within all the provided files are pooled together. Format recognition is automatic. Duplicates are removed (if they have the same name). Duplicates in a single file are only tolerated in FASTA format file, although they will cause sequences to be renamed.
In the above case, the total set of sequences will be made of sequences contained in sequences1.seq, alignment1.aln, alignment2.msf and library.lib, plus the sequences initially gathered by -infile.
2-Turn alignments into libraries
alignment1.aln and alignment2.msf will be read and turned into libraries. Another library will be produced by applying the method lalign_id_pair to the set of sequences previously obtained (1). The final library used for the alignment will be the combination of all this information.
Note as well the following rules:
1-Order: The order in which sequences, methods, alignments and libraries are fed in is irrelevant.
2-Heterogeneity: There is no need for each element (A, S, L) to contain the same sequences.
3-No Duplicate: Each file should contain only one copy of each sequence. Duplicates are only allowed in FASTA files but will cause the sequences to be renamed.
4-Reconciliation: If two files (for instance two alignments) contain different versions of the same sequence due to an indel, a new sequence will be reconstructed and used instead:
aln 1:hgab1 AAAAABAAAAA
aln 2:hgab1 AAAAAAAAAACCC
will cause the program to reconstruct and use the following sequence
hgab1 AAAAABAAAAACCC
This can be useful if you are trying to combine several runs of blast, or structural information where residues may have been deleted. However substitutions are forbidden. If two sequences with the same name cannot be merged, they will cause the program to exit with an information message.
5-Methods: The method describer can either be built in (See ### for a list of all the available methods) or be a file describing the method to be used. The exact syntax is provided in part 4 of this manual.
6-Substitution Matrices: If the method is a substitution matrix (X) then no other type of information should be provided. For instance:
PROMPT: t_coffee sample_seq1.fasta -in=Xpam250mt -gapopen=-10 -gapext=-1
This command results in a progressive alignment carried out on the sequences in seqfile. The procedure does not use any more the T-Coffee concistency based algorithm, but switches to a standard progressive alignment algorithm (like ClustalW or Pileup) much less accurate. In this context, appropriate gap penalties should be provided. The matrices are in the file source/matrices.h. Add-Hoc matrices can also be provided by the user (see the matrices format section at the end of this manual).
Warning: Xmatrix does not have the same effect as using the -matrix flag. The -matrix defines the matrix that will be used while compiling the library while the Xmatrix defines the matrix used when assembling the final alignment.
Usage: -profile=[<name>,] maximum of 200 profiles.
Default: no default
This flag causes T-Coffee to treat multiple alignments as a single sequences, thus making it possible to make multiple profile alignments. The profile-profile alignment is controlled by -profile_mode and -profile_comparison. When provided with the -in flag, profiles must be preceded with the letter R.
PROMPT: t_coffee –profile sample_aln1.aln,sample_aln2.aln –outfile=profile_aln
PROMPT: t_coffee –in Rsample_aln1.aln,Rsample_aln2.aln,Mslow_pair,Mlalign_id_pair –outfile=profile_aln
Note that when using –template_file, the program will also look for the templates associated with the profiles, even if the profiles have been provided as templates themselves (however it will not look for the template of the profile templates of the profile templates…)
Usage: -profile1=[<name>], one name only
Default: no default
Similar to the previous one and was provided for compatibility with ClustalW.
Usage: -profile1=[<name>], one name only
Default: no default
Similar to the previous one and was provided for compatibility with ClustalW.
Usage: -lalign_n_top=<Integer>
Default: -lalign_n_top=10
Number of alignment reported by the local method (lalign).
Unsuported
Usage: -lib_list=<filename>
Default:unset
Use this flag if you do not want the library computation to take into account all the possible pairs in your dataset. For instance
Format:
2 Name1 name2
2 Name1 name4
3 Name1 Name2 Name3…
(the line 3 would be used by a multiple alignment method).
Usage: -do_normalise=<0 or a positive value>
Default:-do_normalise=1000
Development Only
When using a value different from 0, this flag sets the score of the highest scoring pair to 1000.
Usage: -extend=<0,1 or a positive value>
Default:-extend=1
Development Only
When turned on, this flag indicates that the library extension should be carried out when performing the multiple alignment. If -extend =0, the extension is not made, if it is set to 1, the extension is made on all the pairs in the library. If the extension is set to another positive value, the extension is only carried out on pairs having a weight value superior to the specified limit.
Usage: -extend=<string>
Default:-extend=very_fast_triplet
Warning: Development Only
Controls the algorithm for matrix extension. Available modes include:
relative_triplet Unsupported
g_coffee Unsupported
g_coffee_quadruplets Unsupported
fast_triplet Fast triplet extension
very_fast_triplet slow triplet extension, limited to the -max_n_pair best sequence pairs when aligning two profiles
slow_triplet Exhaustive use of all the triplets
mixt Unsupported
quadruplet Unsupported
test Unsupported
matrix Use
of the matrix -matrix
fast_matrix Use of the matrix -matrix. Profiles are turned into consensus
Usage: -max_n_pair=<integer>
Default:-extend=10
Development Only
Controls the number of pairs considered by the -extend_mode=very_fast_triplet. Setting it to 0 forces all the pairs to be considered (equivalent to -extend_mode=slow_triplet).
Usage: Unsupported
Usage: Unsupported
Usage: Unsupported
Usage: Unsupported
Usage: Flag -do_self
Default: No
This flag causes the extension to carried out within the sequences (as opposed to between sequences). This is necessary when looking for internal repeats with Mocca.
Usage: Unsupported
Usage: -weight=<winsimN, sim or sim_<matrix_name or matrix_file> or <integer value>
Default: -weight=sim
Weight defines the way alignments are weighted when turned into a library. Overweighting can be obtained with the OW<X> weight mode.
winsimN indicates that the weight assigned to a given pair will be equal to the percent identity within a window of 2N+1 length centered on that pair. For instance winsim10 defines a window of 10 residues around the pair being considered. This gives its own weight to each residue in the output library. In our hands, this type of weighting scheme has not provided any significant improvement over the standard sim value.
PROMPT: t_coffee sample_seq1.fasta -weight=winsim10 –out_lib=test.tc_lib
sim indicates that the weight equals the average identity within the sequences containing the matched residues.
OW<X> Will
cause the sim weight to be multiplied by X
sim_matrix_name indicates the average identity with two residues regarded as identical when their substitution value is positive. The valid matrices names are in matrices.h (pam250mt) .Matrices not found in this header are considered to be filenames. See the format section for matrices. For instance, -weight=sim_pam250mt indicates that the grouping used for similarity will be the set of classes with positive substitutions.
PROMPT: t_coffee sample_seq1.fasta -weight=winsim10 –out_lib=test.tc_lib
Other groups include
sim_clustalw_col ( categories of clustalw marked with :)
sim_clustalw_dot ( categories of clustalw marked with .)
Value indicates that all the pairs found in the alignments must be given the same weight equal to value. This is useful when the alignment one wishes to turn into a library must be given a pre-specified score (for instance if they come from a structure super-imposition program). Value is an integer:
PROMPT: t_coffee sample_seq1.fasta -weight=1000 –out_lib=test.tc_lib
Usage: -distance_matrix_mode=<slow, fast, very_fast>
Default: very_fast
This flag indicates the method used for computing the distance matrix (distance between every pair of sequences) required for the computation of the dendrogram.
Slow The chosen dp_mode using the extended library,
fast: The fasta dp_mode using the extended library.
very_fast The fasta dp_mode using blosum62mt.
ktup Ktup matching (Muscle kind)
aln Read the distances on a precomputed MSA
Usage: -quicktree
Description: Causes T-Coffee to compute a fast approximate guide tree
This flag is kept for compatibility with ClustalW. It indicates that:
PROMPT: t_coffee sample_seq1.fasta –distance_matrix_mode=very_fast
PROMPT: t_coffee sample_seq1.fasta –quicktree
Controlling
Alignment Computation
Most parameters in this section refer to the alignment mode fasta_pair_wise and cfatsa_pair_wise. When using these alignment modes, things proceed as follow:
1-Sequences are recoded using a degenerated alphabet provided with <-sim_matrix>
2-Recoded sequences are then hashed into ktuples of size <-ktup>
3-Dynamic programming runs on the <-ndiag> best diagonals whose score is higher than <-diag_threshold>, the way diagonals are scored is controlled via <-diag_mode> .
4-The Dynamic computation is made to optimize either the library scoring scheme (as defined by the -in flag) or a substitution matrix as provided via the -matrix flag. The penalty scheme is defined by -gapopen and -gapext. If -gapopen is undefined, the value defined in -cosmetic_penalty is used instead.
5-Terminal gaps are scored according to -tg_mode
Usage: -dp_mode=<string>
Default: -dp_mode=cfasta_fair_wise
This flag indicates the type of dynamic programming used by the program:
PROMPT: t_coffee sample_seq1.fasta –dp_mode myers_miller_pair_wise
gotoh_pair_wise: implementation of the gotoh algorithm (quadratic in memory and time)
myers_miller_pair_wise:
implementation of the Myers and Miller dynamic programming algorithm (
quadratic in time and linear in space). This algorithm is recommended for very
long sequences. It is about 2 times slower than gotoh and only accepts tg_mode=1or 2 (i.e. gaps penalized for
opening).
fasta_pair_wise: implementation of the fasta algorithm.
The sequence is hashed, looking for ktuples
words. Dynamic programming is only carried out on the ndiag best scoring diagonals. This is much faster but less accurate
than the two previous. This mode is controlled by the parameters -ktuple,
-diag_mode and -ndiag
cfasta_pair_wise:
c stands for checked. It is the same algorithm. The dynamic programming is made
on the ndiag best diagonals, and then
on the 2*ndiags, and so on until the scores converge. Complexity will depend on
the level of divergence of the sequences, but will usually be L*log(L), with an
accuracy comparable to the two first mode ( this was checked on BaliBase). This
mode is controlled by the parameters -ktuple, -diag_mode and –ndiag
Note: Users may find by
looking into the code that other modes with fancy names exists
(viterby_pair_wise…) Unless mentioned in this documentation, these modes are
not supported.
Usage: -ktuple=<value>
Default: -ktuple=1 or 2
Indicates the ktuple size for cfasta_pair_wise dp_mode and fasta_pair_wise. It is set to 1 for proteins, and 2 for DNA. The alphabet used for protein can be a degenerated version, set with -sim_matrix..
Usage: -ndiag=<value>
Default: -ndiag=0
Indicates the number of diagonals used by the fasta_pair_wise algorithm (cf -dp_mode). When -ndiag=0, n_diag=Log (length of the smallest sequence)+1.
When –ndiag and
–diag_threshold are set, diagonals are selected if and only if they
fulfill both conditions.
Usage: -diag_mode=<value>
Default: -diag_mode=0
Indicates the manner in which diagonals are scored during the fasta hashing.
0: indicates that the score of a diagonal is equal to the sum of the scores of the exact matches it contains.
1 indicates that this score is set equal to the score of the best uninterrupted segment (useful when dealing with fragments of sequences).
Usage: -diag_threshold=<value>
Default: -diag_threshold=0
Sets the value of the threshold when selecting diagonals.
0: indicates that –ndiag should be used to select the diagonals (cf –ndiag section).
Usage: -sim_matrix=<string>
Default: -sim_matrix=vasiliky
Indicates the manner in which the amino acid alphabet is degenerated when hashing in the fasta_pairwise dynamic programming. Standard ClustalW matrices are all valid. They are used to define groups of amino acids having positive substitution values. In T-Coffee, the default is a 13 letter grouping named Vasiliky, with residues grouped as follows:
rk, de, qh, vilm, fy (other residues kept alone).
This alphabet is set with the flag -sim_matrix=vasiliky. In order to keep the alphabet non degenerated, -sim_matrix=idmat can be used to retain the standard alphabet.
Usage: -matrix=<blosum62mt>
Default: -matrix=blosum62mt
The usage of this flag has been modified from previous versions, due to frequent mistakes in its usage. This flag sets the matrix that will be used by alignment methods within t_coffee (slow_pair, lalign_id_pair). It does not affect external methods (like clustal_pair, clustal_aln…).
Users can also provide their own matrices, using the matrix format described in the appendix.
Usage: -nomatch=<positive value>
Default: -nomatch=0
Indicates the penalty to associate with a match. When using a library, all matches are positive or equal to 0. Matches equal to 0 are unsupported by the library but non-penalized. Setting nomatch to a non-negative value makes it possible to penalize these null matches and prevent unrelated sequences from being aligned (this can be useful when the alignments are meant to be used for structural modeling).
Usage: -gapopen=<negative value>
Default: -gapopen=0
Indicates the penalty applied for opening a gap. The penalty must be negative. If no value is provided when using a substitution matrix, a value will be automatically computed.
Here are some guidelines regarding the tuning of gapopen and gapext. In T-Coffee matches get a score between 0 (match) and 1000 (match perfectly consistent with the library). The default cosmetic penalty is set to -50 (5% of a perfect match). If you want to tune -gapoen and see a strong effect, you should therefore consider values between 0 and -1000.
Usage: -gapext=<negative value>
Default: -gapext=0
Indicates the penalty applied for extending a gap (cf -gapopen)
Unsupported
Unsupported
Usage: -cosmetic_penalty=<negative value>
Default: -cosmetic_penalty=-50
Indicates the penalty applied for opening a gap. This penalty is set to a very low value. It will only have an influence on the portions of the alignment that are unalignable. It will not make them more correct, but only more pleasing to the eye ( i.e. Avoid stretches of lonely residues).
The cosmetic penalty is automatically turned off if a substitution matrix is used rather than a library.
Usage: -tg_mode=<0, 1, or 2>
Default: -tg_mode=1
0: terminal gaps penalized with -gapopen + -gapext*len
1: terminal gaps penalized with a -gapext*len
2: terminal gaps unpenalized.
Usage: -seq_weight=<t_coffee or <file_name>>
Default: -seq_weight=t_coffee
These are the individual weights assigned to each sequence. The t_coffee weights try to compensate the bias in consistency caused by redundancy in the sequences.
sim(A,B)=%similarity between A and B, between 0 and 1.
weight(A)=1/sum(sim(A,X)^3)
Weights are normalized so that their sum equals the number of sequences. They are applied onto the primary library in the following manner:
res_score(Ax,By)=Min(weight(A), weight(B))*res_score(Ax, By)
These are very simple weights. Their main goal is to prevent a single sequence present in many copies to dominate the alignment.
Note: The library output by
-out_lib is the un-weighted
library.
Note: Weights can be output
using the -outseqweight flag.
Note: You can use your own
weights (see the format section).
Usage: -msa_mode=<tree,graph,precomputed>
Default: -evaluate_mode=tree
Unsupported
Usage: -one2all=<name>
Default: not used
Will generate a one to all library with respect to the specified sequence and will then align all the sequences in turn to that sequence, in a sequence determined by the order in which the sequences were provided.
–profile_comparison =profile, the MSAs provided via –profile are vectorized and the function specified by –profile_comparison is used to make profile profile alignments. In that case, the complexity is NL^2
-profile_comparison
Usage: -profile_mode=<fullN,profile>
Default: -profile_mode=full50
The profile mode flag controls the multiple profile alignments in T-Coffee. There are two instances where t_coffee can make multiple profile alignments:
1-When N, the number of sequences is higher than –maxnseq, the program switches to its multiple profile alignment mode (t_coffee_dpa).
2-When MSAs are provided via the –profile flag or via –profile1 and –profile2.
In these situations, the –profile_mode value influences the alignment computation, these values are:
–profile_comparison =profile, the MSAs provided via –profile are vectorized and the function specified by –profile_comparison is used to make profile profile alignments. In that case, the complexity is NL^2
-profile_comparison=fullN, N is an integer value that can omitted. Full indicates that given two profiles, the alignment will be based on a library that includes every possible pair of sequences between the two profiles. If N is set, then the library will be restricted to the N most similar pairs of sequences between the two profiles, as judged from a measure made on a pairwise alignment of these two profiles.
Usage: -profile_mode=<cw_profile_profile, muscle_profile_profile, multi_channel>
Default: -profile_mode=cw_profile_profile
When –profile_comparison=profile, this flag selects a profile scoring function.
Usage: -clean_aln
Default:-clean_aln
This flag causes T-Coffee to post-process the multiple alignment. Residues that have a reliability score smaller or equal to -clean_threshold (as given by an evaluation that uses -clean_evaluate_mode) are realigned to the rest of the alignment. Residues with a score higher than the threshold constitute a rigid framework that cannot be altered.
The cleaning algorithm is greedy. It starts from the top left segment of low constituency residues and works its way left to right, top to bottom along the alignment. You can require this operation to be carried out for several cycles using the -clean_iterations flag.
The rationale behind this operation is mostly cosmetic. In order to ensure a decent looking alignment, the gop is set to -20 and the gep to -1. There is no penalty for terminal gaps, and the matrix is blosum62mt.
Note: Gaps are always
considered to have a reliability score of 0.
Note: The use of the cleaning option can
result in memory overflow when aligning large sequences,
Usage: -clean_threshold=<0-9>
Default:-clean_aln=1
See -clean_aln for details.
Usage: -clean_iteration=<value between 1 and >
Default:-clean_iteration=1
See -clean_aln for details.
Usage: -clean_iteration=<evaluation_mode >
Default:-clean_iteration=t_coffee_non_extended
Indicates the mode used for the evaluation that will indicate the segments that should be realigned. See -evaluation_mode for the list of accepted modes.
Usage: -iterate=<integer>
Default: -iterate=0
Sequences are extracted in turn and realigned to the MSA. If iterate is set to -1, each sequence is realigned, otherwise the number of iterations is set by –iterate.
These parameters are used by T-Coffee when running expresso, accurate and psicoffee
Usage:
-blast_server= EBI, NCBI or LOCAL_BLAST
Default: EBI
Defines whih way BLAST
will be used
Usage: -prot_min_sim= <percent_id>
Default: 40
Minimum id for inclusion
of a sequence in a psi-blast profile
Usage: -prot_max_sim= <percent_id>
Default: 90
Maximum id for inclusion
of a sequence in a psi-blast profile.
Usage: -prot_min_cov= <percent>
Default: 40
Minimum coverage for
inclusion of a sequence in a psi-blast profile
Usage: -protein_db= <BLAST database>
Default: nr
Database used for construction of psi-blast profiles
Usage: -pdb_min_sim= <percent_id>
Default: 35
Minimum id for a PDB
template to be selected by expresso
Usage: -pdb_max_sim= <percent_id>
Default: 100
Maximum id for a PDB
template to be selected by expresso
Usage: -pdb_min_cov= <percent>
Default: 50
Minimum coverage for a
PDB template to be selected by expresso.
Usage: -protein_db= <BLAST database>
Default: pdb
Database for PDB template to be selected by expresso.
Usage: -pdb_type= d,n,m,dnm,dn
Default: d
d: diffraction
n: NMR
m: model
Usage: -multi_core=
templates_jobs_relax_msa
Default: 0
template: fetch the templates in a parallel way
jobs: compute the library
relax: extend the library in a parallel way
msa: compute the msa in a parallel way
Specifies that the steps of T-Coffee that should be multi threaded. by default all relevant steps are parallelized.
PROMPT: t_coffee sample_seq2.fasta -multi_core jobs
In order to prevent the use of the parallel mode it is possible to use:
PROMPT: t_coffee sample_seq2.fasta -multi_core no
Usage:
-n_core= <number of cores>
Default: 0
Default indicates that all cores will be used, as indicated by the environment via:
PROMPT: t_coffee sample_seq2.fasta -multi_core jobs
Usage:
deprecated
Usage: -ulimit=<value>
Default: -ulimit=0
Specifies the upper limit of memory usage (in Megabytes). Processes exceeding this limit will automatically exit. A value 0 indicates that no limit applies.
Usage: -maxlen=<value, 0=nolimit>
Default: -maxlen=1000
Indicates the maximum length of the sequences.
Usage: -maxnseq=<value, 0=nolimit>
Default: -maxnseq=50
Indicates the maximum number of sequences before triggering the use of t_coffee_dpa.
Usage: -dpa_master_aln=<File, method>
Default: -dpa_master_aln=NO
When using dpa, t_coffee needs a seed alignment that can be computed using any appropriate method. By default, t_coffee computes a fast approximate alignment.
A pre-alignment can be provided through this flag, as well as any program using the following syntax:
your_script –in <fasta_file> -out <file_name>
Usage: -dpa_maxnseq=<integer value>
Default: -dpa_maxnseq=30
Maximum number of sequences aligned simultaneously when DPA is ran. Given the tree computed from the master alignment, a node is sent to computation if it controls more than –dpa_maxnseq OR if it controls a pair of sequences having less than –dpa_min_score2 percent ID.
Usage: -dpa_min_score1=<integer value>
Default: -dpa_min_score1=95
Threshold for not realigning the sequences within the master alignment. Given this alignment and the associated tree, sequences below a node are not realigned if none of them has less than –dpa_min_score1 % identity.
Usage: -dpa_min_score2
Default: -dpa_min_score2
Maximum number of sequences aligned simultaneously when DPA is ran. Given the tree computed from the master alignment, a node is sent to computation if it controls more than –dpa_maxnseq OR if it controls a pair of sequences having less than –dpa_min_score2 percent ID.
Usage: -dpa_tree=<filename>
Default: -unset
Guide tree used in DPA. This is a newick tree where the distance associated with each node is set to the minimum pairwise distance among all considered sequences.
Usage: -mode=3dcoffee
Default: turned off
Runs t_coffee with the 3dcoffee mode (cf next section).
Usage: -check_pdb_status
Default: turned off
Forces t_coffee to run extract_from_pdb to check the pdb status of each sequence. This can considerably slow down the program.
It is possible to use t_coffee to compute multiple structural alignments. To do so, ensure that you have the sap program installed.
PROMPT: t_coffee –pdb=struc1.pdb,struc2.pdb,struc3.pdb -method sap_pair
Will combine the pairwise alignments produced by SAP. There are currently four methods that can be interfaced with t_coffee:
sap_pair: that uses the sap algorithm
align_pdb: uses a t_coffee implementation of sap, not as accurate.
tmaliagn_pair (http://zhang.bioinformatics.ku.edu/TM-align/)
mustang_pair (http://www.cs.mu.oz.au/~arun/mustang)
When providing a PDB file, the computation is only carried out on the first chain of this file. If your original file contains several chain, you should extract the chain you want to work on. You can use t_coffee –other_pg extract_from_pdb or any pdb handling program.
If you are working with public PDB files, you can use the PDB identifier and specify the chain by adding its index to the identifier (i.e. 1pdbC). If your structure is an NMR structure, you are advised to provide the program with one structure only.
If you wish to align only a portion of the structure, you should extract it yourself from the pdb file, using t_coffee –other_pg extract_from_pdb or any pdb handling program.
You can provide t_coffee with a mixture of sequences and structure. In this case, you should use the special mode:
PROMPT: t_coffee –mode 3dcoffee –seq 3d_sample3.fasta -template_file template_file.template
Usage: -template_file =
<filename,
SCRIPT_scriptame,
SELF_TAG
SEQFILE_TAG_filename,
no>
Default: no
This flag instructs t_coffee on the templates that will be used when combining several types of information. For instance, when using structural information, this file will indicate the structural template that corresponds to your sequences. The identifier T indicates that the file should be a FASTA like file, formatted as follows. There are several ways to pass the templates:
Predefined Modes
EXPRESSO: will use the EBI server to find _P_ templates
PSIBLAST: will use the EBI sever to find profiles
File name
This file contains the sequence/template association it uses a FASTA-like format, as follows:
><sequence name> _P_ <pdb template>
><sequence name> _G_ <gene template>
><sequence name> _R_ <MSA template>
><sequence name> _F_ <RNA Secondary Structure>
><sequence name> _T_ <Transmembrane Secondary Structure>
><sequence name> _E_ <Protein Secondary Structure>
Each template will be used in place of the sequence with the appropriate method. For instance, structural templates will be aligned with sap_pair and the information thus generated will be transferred onto the alignment.
Note the following rule:
-Each sequence can have one template of each type (structural, genomics…)
-Each sequence can only have one template of a given type
-Several sequences can share the same template
-All the sequences do not need to have a template
The type of template on which a method works is declared with the SEQ_TYPE parameter in the method configuration file:
SEQ_TYPE S: a method that uses sequences
SEQ_TYPE PS: a pairwise method that aligns sequences and structures
SEQ_TYPE P: a method that aligns structures (sap for instance)
There are 4 tags identifying the template type:
_P_ Structural templates: a pdb identifier OR a pdb file
_G_ Genomic templates: a protein sequence where boundary amino-acid have been recoded with ( o:0, i:1, j:2)
_R_ Profile Templates: a file containing a multiple sequence alignment
_F_ RNA secondary Structures
More than one template file can be provided. There is no need to have one template for every sequence in the dataset.
_P_, _G_, and _R_ are known as template TAGS
2-SCRIPT_<scriptname>
Indicates that filename is a script that will be used to generate a valid template file. The script will run on a file containing all your sequences using the following syntax:
scriptname –infile=<your sequences> -outfile=<template_file>
It is also possible to pass some parameters, use @ as a separator and # in place of the = sign. For instance, if you want to call the a script named blast.pl with the foloowing parameters;
blast.pl -db=pdb -dir=/local/test
Use
SCRIPT_blast.pl@db#pdb@dir#/local/test
Bear in mind that the input output flags will then be concatenated to this command line so that t_coffee ends up calling the program using the following system call:
blast.pl -db=pdb -dir=/local/test -infile=<some tmp file> -outfile=<another tmp file>
3-SELF_TAG
TAG can take the value of any of the known TAGS (_S_, _G_, _P_). SELF indicates that the original name of the sequence will be used to fetch the template:
PROMPT: t_coffee 3d_sample2.fasta –template_file SELF_P_
The previous command will work because the sequences in 3d_sample3 are named
4-SEQFILE_TAG_filename
Use this flag if your templates are in filename, and are named according to the sequences. For instance, if your protein sequences have been recoded with Exon/Intron information, you should have the recoded sequences names according to the original:
SEQFILE_G_recodedprotein.fasta
Usage: -struc_to_use=<struc1, struc2…>
Default: -struc_to_use=NULL
Restricts the 3Dcoffee to a set of pre-defined structures.
It is possible to compute multiple local alignments, using the moca routine. MOCA is a routine that allows extracting all the local alignments that show some similarity with another predefined fragment.
'mocca' is a perl script that calls t-coffee and provides it with the appropriate parameters.
Usage: -domain
Default: not set
This flag indicates that t_coffee will run using the domain mode. All the sequences will be concatenated, and the resulting sequence will be compared to itself using lalign_rs_s_pair mode (lalign of the sequence against itself using keeping the lalign raw score). This step is the most computer intensive, and it is advisable to save the resulting file.
PROMPT: t_coffee -in Ssample_seq1.fasta,Mlalign_rs_s_pair -out_lib=sample_lib1.mocca_lib -domain -start=100 -len=50
This instruction will use the fragment 100-150 on the concatenated sequences, as a template for the extracted repeats. The extraction will only be made once. The library will be placed in the file <lib name>.
If you want, you can test other coordinates for the repeat, such as
PROMPT: t_coffee -in sample_lib1.mocca_lib -domain -start=100 -len=60
This run will use the fragment 100-160, and will be much faster because it does not need to re-compute the lalign library.
Usage: -start=<int value>
Default: not set
This flag indicates the starting position of the portion of sequence that will be used as a template for the repeat extraction. The value assumes that all the sequences have been concatenated, and is given on the resulting sequence.
Usage: -len=<int value>
Default: not set
This flag indicates the length of the portion of sequence that will be used as a template.
Usage: -scale=<int value>
Default: -scale=-100
This flag indicates the value of the threshold for extracting the repeats. The actual threshold is equal to:
motif_len*scale
Increase the scale óIncrease sensitivity ó More alignments( i.e. -50).
-domain_interactive [Examples]
Usage: -domain_interactive
Default: unset
Launches an interactive mocca session.
PROMPT: t_coffee -in Lsample_lib3.tc_lib,Mlalign_rs_s_pair -domain -start=100 -len=60
TOLB_ECOLI_212_26 211 SKLAYVTFESGR--SALVIQTLANGAVRQV-ASFPRHNGAPAFSPDGSKLAFA
TOLB_ECOLI_165_218 164 TRIAYVVQTNGGQFPYELRVSDYDGYNQFVVHRSPQPLMSPAWSPDGSKLAYV
TOLB_ECOLI_256_306 255 SKLAFALSKTGS--LNLYVMDLASGQIRQV-TDGRSNNTEPTWFPDSQNLAFT
TOLB_ECOLI_307_350 306 -------DQAGR--PQVYKVNINGGAPQRI-TWEGSQNQDADVSSDGKFMVMV
TOLB_ECOLI_351_393 350 -------SNGGQ--QHIAKQDLATGGV-QV-LSSTFLDETPSLAPNGTMVIYS
1 * * : . .:. :
MENU: Type Letter Flag[number] and Return: ex |10
|x -->Set the START to x
>x -->Set the LEN to x
Cx -->Set the sCale to x
Sname -->Save the Alignment
Bx -->Save Goes back x it
return -->Compute the Alignment
X -->eXit
[ITERATION 1] [START=211] [LEN= 50] [SCALE=-100] YOUR CHOICE:
For instance, to set the length of the domain to 40, type:
[ITERATION 1] [START=211] [LEN= 50] [SCALE=-100] YOUR CHOICE:>40[return]
[return]
Which will generate:
TOLB_ECOLI_212_252 211 SKLAYVTFESGRSALVIQTLANGAVRQVASFPRHNGAPAF 251
TOLB_ECOLI_256_296 255 SKLAFALSKTGSLNLYVMDLASGQIRQVTDGRSNNTEPTW 295
TOLB_ECOLI_300_340 299 QNLAFTSDQAGRPQVYKVNINGGAPQRITWEGSQNQDADV 339
TOLB_ECOLI_344_383 343 KFMVMVSSNGGQQHIAKQDLATGGV-QVLSSTFLDETPSL 382
TOLB_ECOLI_387_427 386 TMVIYSSSQGMGSVLNLVSTDGRFKARLPATDGQVKFPAW 426
1 : : : :: . 40
MENU: Type Letter Flag[number] and Return: ex |10
|x -->Set the START to x
>x -->Set the LEN to x
Cx -->Set the sCale to x
Sname -->Save the Alignment
Bx -->Save Goes back x it
return -->Compute the Alignment
X -->eXit
[ITERATION 3] [START=211] [LEN= 40] [SCALE=-100] YOUR CHOICE:
If you want to indicate the coordinates, relative to a specific sequence, type:
|<seq_name>:start
Type S<your name> to save the current alignment, and extract a new motif.
Type X when you are done.
Conventions Regarding Filenames
stdout, stderr, stdin, no, /dev/null are valid filenames. They cause the corresponding file to be output in stderr or stdout, for an input file, stdin causes the program to requests the corresponding file through pipe. No causes a suppression of the output, as does /dev/null.
Identifying the Output files automatically
In the t_coffee output, each output appears in a line:
##### FILENAME <name> TYPE <Type> FORMAT <Format>
Usage: -no_warning
Default: Switched off
Suppresseswarning
output.
Usage: -outfile=<out_aln file,default,no>
DefauWord did not find any entries for your table of contents.lt:-outfile=default
Indicates the name of the alignment output by
t_coffee. If the default is used, the alignment is named <your sequences>.aln
Usage: -output=<format1,format2,...>
Default:-output=clustalw
Indicates the format used for outputting the -outfile.
Supported formats are:
clustalw_aln, clustalw : ClustalW format.