ࡱ>  SUJKLMNOPQR` Ǥbjbj ZlNNNNNNNDN$@LNt\ :..Ҿ.C4G EtGtGtGtGtGtGt$wh,zZktND$D$D$ktNN.Ҿt]]]D$)N.NҾEt]D$Et]] hNN9m. z@NT l,rt0tlPzl[BzX9mzN9mP ]m!!Sktkt\dtD$D$D$D$Nyy@ N@ NNNNNNNNN Technical Centre National De LA Recherche scientifique (France) CeNTRO De REGULACIO GENOMICA (SPAIN) Cdric Notredame www.tcoffee.org T-Coffee: Technical Documentation T-Coffee Technical Documentation (Version 5.31, December 2007) www.tcoffee.org T-Coffee 3D-Coffee M-Coffee R-Coffee APDB and iRMSD ( Cdric Notredame and Centre National de la Recherche Scientifique, France  TOC \h \z \t "Heading 1;2;Heading 2;3;Heading 3;4;Chapter Title;1;flag;4;FAQ;4"  HYPERLINK \l "_Toc138137779" License and Terms of Use  PAGEREF _Toc138137779 \h 5  HYPERLINK \l "_Toc138137780" T-Coffee is distributed under the Gnu Public License  PAGEREF _Toc138137780 \h 5  HYPERLINK \l "_Toc138137781" T-Coffee code can be re-used freely  PAGEREF _Toc138137781 \h 5  HYPERLINK \l "_Toc138137782" T-Coffee can be incorporated in any pipeline: Plug-in/Plug-out  PAGEREF _Toc138137782 \h 5  HYPERLINK \l "_Toc138137783" Addresses and Contacts  PAGEREF _Toc138137783 \h 6  HYPERLINK \l "_Toc138137784" Contributors  PAGEREF _Toc138137784 \h 6  HYPERLINK \l "_Toc138137785" Addresses  PAGEREF _Toc138137785 \h 6  HYPERLINK \l "_Toc138137786" Citations  PAGEREF _Toc138137786 \h 8  HYPERLINK \l "_Toc138137787" T-Coffee  PAGEREF _Toc138137787 \h 8  HYPERLINK \l "_Toc138137788" Mocca  PAGEREF _Toc138137788 \h 9  HYPERLINK \l "_Toc138137789" CORE  PAGEREF _Toc138137789 \h 10  HYPERLINK \l "_Toc138137790" Other Contributions  PAGEREF _Toc138137790 \h 10  HYPERLINK \l "_Toc138137791" Bug Reports and Feedback  PAGEREF _Toc138137791 \h 10  HYPERLINK \l "_Toc138137792" Installation  PAGEREF _Toc138137792 \h 11  HYPERLINK \l "_Toc138137793" Standard Installation of T-Coffee  PAGEREF _Toc138137793 \h 11  HYPERLINK \l "_Toc138137794" Installation of M-Coffee  PAGEREF _Toc138137794 \h 12  HYPERLINK \l "_Toc138137795" Installation of APDB and iRMSD  PAGEREF _Toc138137795 \h 13  HYPERLINK \l "_Toc138137796" Installation of seq_reformat  PAGEREF _Toc138137796 \h 13  HYPERLINK \l "_Toc138137797" Installation of extract_from_pdb  PAGEREF _Toc138137797 \h 13  HYPERLINK \l "_Toc138137798" Installation of 3D-Coffee  PAGEREF _Toc138137798 \h 13  HYPERLINK \l "_Toc138137799" Quick Start  PAGEREF _Toc138137799 \h 15  HYPERLINK \l "_Toc138137800" T-COFFEE  PAGEREF _Toc138137800 \h 15  HYPERLINK \l "_Toc138137801" M-Coffee  PAGEREF _Toc138137801 \h 15  HYPERLINK \l "_Toc138137802" iRMSD and APDB  PAGEREF _Toc138137802 \h 16  HYPERLINK \l "_Toc138137803" MOCCA  PAGEREF _Toc138137803 \h 16  HYPERLINK \l "_Toc138137804" Recent Modifications  PAGEREF _Toc138137804 \h 17  HYPERLINK \l "_Toc138137805" Reference Manual  PAGEREF _Toc138137805 \h 18  HYPERLINK \l "_Toc138137806" Environment Variables  PAGEREF _Toc138137806 \h 18  HYPERLINK \l "_Toc138137807" DIR_4_TCOFFEE  PAGEREF _Toc138137807 \h 18  HYPERLINK \l "_Toc138137808" TMP_4_TCOFFEE  PAGEREF _Toc138137808 \h 18  HYPERLINK \l "_Toc138137809" CACHE_4_TCOFFEE  PAGEREF _Toc138137809 \h 18  HYPERLINK \l "_Toc138137810" NO_ERROR_REPORT_4_TCOFFEE  PAGEREF _Toc138137810 \h 18  HYPERLINK \l "_Toc138137811" PDB_DIR  PAGEREF _Toc138137811 \h 19  HYPERLINK \l "_Toc138137812" NO_WARNING_4_TCOFFEE  PAGEREF _Toc138137812 \h 19  HYPERLINK \l "_Toc138137813" Well Behaved Parameters  PAGEREF _Toc138137813 \h 19  HYPERLINK \l "_Toc138137814" Separation  PAGEREF _Toc138137814 \h 19  HYPERLINK \l "_Toc138137815" Posix  PAGEREF _Toc138137815 \h 19  HYPERLINK \l "_Toc138137816" Entering the right parameters  PAGEREF _Toc138137816 \h 19  HYPERLINK \l "_Toc138137817" Parameters Syntax  PAGEREF _Toc138137817 \h 20  HYPERLINK \l "_Toc138137818" No Flag  PAGEREF _Toc138137818 \h 20  HYPERLINK \l "_Toc138137819" -parameters  PAGEREF _Toc138137819 \h 20  HYPERLINK \l "_Toc138137820" -t_coffee_defaults  PAGEREF _Toc138137820 \h 20  HYPERLINK \l "_Toc138137821" -special_mode  PAGEREF _Toc138137821 \h 21  HYPERLINK \l "_Toc138137822" -score [Deprecated]  PAGEREF _Toc138137822 \h 21  HYPERLINK \l "_Toc138137823" -evaluate  PAGEREF _Toc138137823 \h 21  HYPERLINK \l "_Toc138137824" -convert [cw]  PAGEREF _Toc138137824 \h 21  HYPERLINK \l "_Toc138137825" -do_align [cw]  PAGEREF _Toc138137825 \h 22  HYPERLINK \l "_Toc138137826" Special Parameters  PAGEREF _Toc138137826 \h 22  HYPERLINK \l "_Toc138137827" -version  PAGEREF _Toc138137827 \h 22  HYPERLINK \l "_Toc138137828" -check_configuration  PAGEREF _Toc138137828 \h 22  HYPERLINK \l "_Toc138137829" -cache  PAGEREF _Toc138137829 \h 22  HYPERLINK \l "_Toc138137830" -update  PAGEREF _Toc138137830 \h 22  HYPERLINK \l "_Toc138137831" -full_log  PAGEREF _Toc138137831 \h 22  HYPERLINK \l "_Toc138137832" -other_pg  PAGEREF _Toc138137832 \h 22  HYPERLINK \l "_Toc138137833" Input  PAGEREF _Toc138137833 \h 23  HYPERLINK \l "_Toc138137834" Sequence Input  PAGEREF _Toc138137834 \h 23  HYPERLINK \l "_Toc138137835" -infile [cw]  PAGEREF _Toc138137835 \h 23  HYPERLINK \l "_Toc138137836" -in (Cf in from the Method and Library Input section)  PAGEREF _Toc138137836 \h 23  HYPERLINK \l "_Toc138137837" -get_type  PAGEREF _Toc138137837 \h 23  HYPERLINK \l "_Toc138137838" -type [cw]  PAGEREF _Toc138137838 \h 23  HYPERLINK \l "_Toc138137839" -seq  PAGEREF _Toc138137839 \h 23  HYPERLINK \l "_Toc138137840" -seq_source  PAGEREF _Toc138137840 \h 23  HYPERLINK \l "_Toc138137841" Structure Input  PAGEREF _Toc138137841 \h 24  HYPERLINK \l "_Toc138137842" -pdb  PAGEREF _Toc138137842 \h 24  HYPERLINK \l "_Toc138137843" Tree Input  PAGEREF _Toc138137843 \h 24  HYPERLINK \l "_Toc138137844" -usetree  PAGEREF _Toc138137844 \h 24  HYPERLINK \l "_Toc138137845" Structures, Sequences Methods and Library Input via the in Flag  PAGEREF _Toc138137845 \h 24  HYPERLINK \l "_Toc138137846" -in  PAGEREF _Toc138137846 \h 25  HYPERLINK \l "_Toc138137847" Profile Input  PAGEREF _Toc138137847 \h 26  HYPERLINK \l "_Toc138137848" -profile  PAGEREF _Toc138137848 \h 26  HYPERLINK \l "_Toc138137849" -profile1 [cw]  PAGEREF _Toc138137849 \h 26  HYPERLINK \l "_Toc138137850" -profile2 [cw]  PAGEREF _Toc138137850 \h 27  HYPERLINK \l "_Toc138137851" Alignment Computation  PAGEREF _Toc138137851 \h 27  HYPERLINK \l "_Toc138137852" Library Computation: Methods  PAGEREF _Toc138137852 \h 27  HYPERLINK \l "_Toc138137853" -lalign_n_top  PAGEREF _Toc138137853 \h 27  HYPERLINK \l "_Toc138137854" -align_pdb_param_file  PAGEREF _Toc138137854 \h 27  HYPERLINK \l "_Toc138137855" -align_pdb_hasch_mode  PAGEREF _Toc138137855 \h 27  HYPERLINK \l "_Toc138137856" Library Computation: Extension  PAGEREF _Toc138137856 \h 27  HYPERLINK \l "_Toc138137857" -lib_list [Unsupported]  PAGEREF _Toc138137857 \h 27  HYPERLINK \l "_Toc138137858" -do_normalise  PAGEREF _Toc138137858 \h 27  HYPERLINK \l "_Toc138137859" -extend  PAGEREF _Toc138137859 \h 28  HYPERLINK \l "_Toc138137860" -extend_mode  PAGEREF _Toc138137860 \h 28  HYPERLINK \l "_Toc138137861" -max_n_pair  PAGEREF _Toc138137861 \h 28  HYPERLINK \l "_Toc138137862" -seq_name_for_quadruplet  PAGEREF _Toc138137862 \h 28  HYPERLINK \l "_Toc138137863" -compact  PAGEREF _Toc138137863 \h 28  HYPERLINK \l "_Toc138137864" -clean  PAGEREF _Toc138137864 \h 29  HYPERLINK \l "_Toc138137865" -maximise  PAGEREF _Toc138137865 \h 29  HYPERLINK \l "_Toc138137866" -do_self  PAGEREF _Toc138137866 \h 29  HYPERLINK \l "_Toc138137867" -seq_name_for_quadruplet  PAGEREF _Toc138137867 \h 29  HYPERLINK \l "_Toc138137868" -weight  PAGEREF _Toc138137868 \h 29  HYPERLINK \l "_Toc138137869" Tree Computation  PAGEREF _Toc138137869 \h 30  HYPERLINK \l "_Toc138137870" -distance_matrix_mode  PAGEREF _Toc138137870 \h 30  HYPERLINK \l "_Toc138137871" -quicktree [CW]  PAGEREF _Toc138137871 \h 30  HYPERLINK \l "_Toc138137872" Pair-wise Alignment Computation  PAGEREF _Toc138137872 \h 30  HYPERLINK \l "_Toc138137873" -dp_mode  PAGEREF _Toc138137873 \h 31  HYPERLINK \l "_Toc138137874" -ktuple  PAGEREF _Toc138137874 \h 31  HYPERLINK \l "_Toc138137875" -ndiag  PAGEREF _Toc138137875 \h 31  HYPERLINK \l "_Toc138137876" -diag_mode  PAGEREF _Toc138137876 \h 31  HYPERLINK \l "_Toc138137877" -diag_threshold  PAGEREF _Toc138137877 \h 32  HYPERLINK \l "_Toc138137878" -sim_matrix  PAGEREF _Toc138137878 \h 32  HYPERLINK \l "_Toc138137879" -matrix [CW]  PAGEREF _Toc138137879 \h 32  HYPERLINK \l "_Toc138137880" -nomatch  PAGEREF _Toc138137880 \h 32  HYPERLINK \l "_Toc138137881" -gapopen  PAGEREF _Toc138137881 \h 32  HYPERLINK \l "_Toc138137882" -gapext  PAGEREF _Toc138137882 \h 33  HYPERLINK \l "_Toc138137883" -fgapopen  PAGEREF _Toc138137883 \h 33  HYPERLINK \l "_Toc138137884" -fgapext  PAGEREF _Toc138137884 \h 33  HYPERLINK \l "_Toc138137885" -cosmetic_penalty  PAGEREF _Toc138137885 \h 33  HYPERLINK \l "_Toc138137886" -tg_mode  PAGEREF _Toc138137886 \h 33  HYPERLINK \l "_Toc138137887" Weighting Schemes  PAGEREF _Toc138137887 \h 33  HYPERLINK \l "_Toc138137888" -seq_weight  PAGEREF _Toc138137888 \h 33  HYPERLINK \l "_Toc138137889" Multiple Alignment Computation  PAGEREF _Toc138137889 \h 34  HYPERLINK \l "_Toc138137890" -msa_mode  PAGEREF _Toc138137890 \h 34  HYPERLINK \l "_Toc138137891" -profile_comparison  PAGEREF _Toc138137891 \h 34  HYPERLINK \l "_Toc138137892" -profile_mode  PAGEREF _Toc138137892 \h 34  HYPERLINK \l "_Toc138137893" Alignment Post-Processing  PAGEREF _Toc138137893 \h 34  HYPERLINK \l "_Toc138137894" -clean_aln  PAGEREF _Toc138137894 \h 34  HYPERLINK \l "_Toc138137895" -clean_threshold  PAGEREF _Toc138137895 \h 35  HYPERLINK \l "_Toc138137896" -clean_iteration  PAGEREF _Toc138137896 \h 35  HYPERLINK \l "_Toc138137897" -clean_evaluation_mode  PAGEREF _Toc138137897 \h 35  HYPERLINK \l "_Toc138137898" -iterate  PAGEREF _Toc138137898 \h 35  HYPERLINK \l "_Toc138137899" CPU Control  PAGEREF _Toc138137899 \h 35  HYPERLINK \l "_Toc138137900" Multithreading  PAGEREF _Toc138137900 \h 35  HYPERLINK \l "_Toc138137901" -multi_thread [NOT Supported]  PAGEREF _Toc138137901 \h 35  HYPERLINK \l "_Toc138137902" Limits  PAGEREF _Toc138137902 \h 36  HYPERLINK \l "_Toc138137903" -mem_mode  PAGEREF _Toc138137903 \h 36  HYPERLINK \l "_Toc138137904" -ulimit  PAGEREF _Toc138137904 \h 36  HYPERLINK \l "_Toc138137905" -maxlen  PAGEREF _Toc138137905 \h 36  HYPERLINK \l "_Toc138137906" Aligning more than 100 sequences with DPA  PAGEREF _Toc138137906 \h 36  HYPERLINK \l "_Toc138137907" -maxnseq  PAGEREF _Toc138137907 \h 36  HYPERLINK \l "_Toc138137908" -dpa_master_aln  PAGEREF _Toc138137908 \h 36  HYPERLINK \l "_Toc138137909" -dpa_maxnseq  PAGEREF _Toc138137909 \h 36  HYPERLINK \l "_Toc138137910" -dpa_min_score1  PAGEREF _Toc138137910 \h 37  HYPERLINK \l "_Toc138137911" -dpa_min_score2  PAGEREF _Toc138137911 \h 37  HYPERLINK \l "_Toc138137912" -dap_tree [NOT IMPLEMENTED]  PAGEREF _Toc138137912 \h 37  HYPERLINK \l "_Toc138137913" Using Structures  PAGEREF _Toc138137913 \h 37  HYPERLINK \l "_Toc138137914" Generic  PAGEREF _Toc138137914 \h 37  HYPERLINK \l "_Toc138137915" -special_mode  PAGEREF _Toc138137915 \h 37  HYPERLINK \l "_Toc138137916" -check_pdb_status  PAGEREF _Toc138137916 \h 37  HYPERLINK \l "_Toc138137917" 3D Coffee: Using SAP  PAGEREF _Toc138137917 \h 38  HYPERLINK \l "_Toc138137918" Using/finding PDB templates for the Sequences  PAGEREF _Toc138137918 \h 38  HYPERLINK \l "_Toc138137919" -template_file  PAGEREF _Toc138137919 \h 38  HYPERLINK \l "_Toc138137920" -struc_to_use  PAGEREF _Toc138137920 \h 39  HYPERLINK \l "_Toc138137921" Multiple Local Alignments  PAGEREF _Toc138137921 \h 40  HYPERLINK \l "_Toc138137922" -domain/-mocca  PAGEREF _Toc138137922 \h 40  HYPERLINK \l "_Toc138137923" -start  PAGEREF _Toc138137923 \h 40  HYPERLINK \l "_Toc138137924" -len  PAGEREF _Toc138137924 \h 40  HYPERLINK \l "_Toc138137925" -scale  PAGEREF _Toc138137925 \h 41  HYPERLINK \l "_Toc138137926" -domain_interactive [Examples]  PAGEREF _Toc138137926 \h 41  HYPERLINK \l "_Toc138137927" Output Control  PAGEREF _Toc138137927 \h 42  HYPERLINK \l "_Toc138137928" Generic  PAGEREF _Toc138137928 \h 42  HYPERLINK \l "_Toc138137929" Conventions Regarding Filenames  PAGEREF _Toc138137929 \h 42  HYPERLINK \l "_Toc138137930" Identifying the Output files automatically  PAGEREF _Toc138137930 \h 42  HYPERLINK \l "_Toc138137931" -no_warning  PAGEREF _Toc138137931 \h 42  HYPERLINK \l "_Toc138137932" Alignments  PAGEREF _Toc138137932 \h 42  HYPERLINK \l "_Toc138137933" -outfile  PAGEREF _Toc138137933 \h 42  HYPERLINK \l "_Toc138137934" -output  PAGEREF _Toc138137934 \h 42  HYPERLINK \l "_Toc138137935" -outseqweight  PAGEREF _Toc138137935 \h 43  HYPERLINK \l "_Toc138137936" -case  PAGEREF _Toc138137936 \h 43  HYPERLINK \l "_Toc138137937" -cpu  PAGEREF _Toc138137937 \h 43  HYPERLINK \l "_Toc138137938" -outseqweight  PAGEREF _Toc138137938 \h 43  HYPERLINK \l "_Toc138137939" -outorder [cw]  PAGEREF _Toc138137939 \h 44  HYPERLINK \l "_Toc138137940" -inorder [cw]  PAGEREF _Toc138137940 \h 44  HYPERLINK \l "_Toc138137941" -seqnos  PAGEREF _Toc138137941 \h 44  HYPERLINK \l "_Toc138137942" Libraries  PAGEREF _Toc138137942 \h 44  HYPERLINK \l "_Toc138137943" -out_lib  PAGEREF _Toc138137943 \h 44  HYPERLINK \l "_Toc138137944" -lib_only  PAGEREF _Toc138137944 \h 44  HYPERLINK \l "_Toc138137945" Trees  PAGEREF _Toc138137945 \h 45  HYPERLINK \l "_Toc138137946" -newtree  PAGEREF _Toc138137946 \h 45  HYPERLINK \l "_Toc138137947" Reliability Estimation  PAGEREF _Toc138137947 \h 45  HYPERLINK \l "_Toc138137948" CORE Computation  PAGEREF _Toc138137948 \h 45  HYPERLINK \l "_Toc138137949" -evaluate_mode  PAGEREF _Toc138137949 \h 45  HYPERLINK \l "_Toc138137950" Generic Output  PAGEREF _Toc138137950 \h 46  HYPERLINK \l "_Toc138137951" -run_name  PAGEREF _Toc138137951 \h 46  HYPERLINK \l "_Toc138137952" -quiet  PAGEREF _Toc138137952 \h 46  HYPERLINK \l "_Toc138137953" -align [CW]  PAGEREF _Toc138137953 \h 46  HYPERLINK \l "_Toc138137954" APDB/iRMSD Parameters  PAGEREF _Toc138137954 \h 46  HYPERLINK \l "_Toc138137955" -quiet [Same as T-Coffee]  PAGEREF _Toc138137955 \h 46  HYPERLINK \l "_Toc138137956" -run_name [Same as T-Coffee]  PAGEREF _Toc138137956 \h 46  HYPERLINK \l "_Toc138137957" -aln  PAGEREF _Toc138137957 \h 46  HYPERLINK \l "_Toc138137958" -n_excluded_nb  PAGEREF _Toc138137958 \h 47  HYPERLINK \l "_Toc138137959" -maximum_distance  PAGEREF _Toc138137959 \h 47  HYPERLINK \l "_Toc138137960" -similarity_threshold  PAGEREF _Toc138137960 \h 47  HYPERLINK \l "_Toc138137961" -local_mode  PAGEREF _Toc138137961 \h 47  HYPERLINK \l "_Toc138137962" -filter  PAGEREF _Toc138137962 \h 47  HYPERLINK \l "_Toc138137963" -print_rapdb [Unsupported]  PAGEREF _Toc138137963 \h 47  HYPERLINK \l "_Toc138137964" -outfile [Same as T-Coffee]  PAGEREF _Toc138137964 \h 48  HYPERLINK \l "_Toc138137965" -color_mode  PAGEREF _Toc138137965 \h 48  HYPERLINK \l "_Toc138137966" Building a Server  PAGEREF _Toc138137966 \h 49  HYPERLINK \l "_Toc138137967" Environment Variables  PAGEREF _Toc138137967 \h 49  HYPERLINK \l "_Toc138137968" Output of the .dnd file.  PAGEREF _Toc138137968 \h 50  HYPERLINK \l "_Toc138137969" Permissions  PAGEREF _Toc138137969 \h 50  HYPERLINK \l "_Toc138137970" Other Programs  PAGEREF _Toc138137970 \h 50  HYPERLINK \l "_Toc138137971" Formats  PAGEREF _Toc138137971 \h 51  HYPERLINK \l "_Toc138137972" Parameter files  PAGEREF _Toc138137972 \h 51  HYPERLINK \l "_Toc138137973" Sequence Name Handling  PAGEREF _Toc138137973 \h 51  HYPERLINK \l "_Toc138137974" Automatic Format Recognition  PAGEREF _Toc138137974 \h 52  HYPERLINK \l "_Toc138137975" Structures  PAGEREF _Toc138137975 \h 52  HYPERLINK \l "_Toc138137976" Sequences  PAGEREF _Toc138137976 \h 52  HYPERLINK \l "_Toc138137977" Alignments  PAGEREF _Toc138137977 \h 52  HYPERLINK \l "_Toc138137978" Libraries  PAGEREF _Toc138137978 \h 53  HYPERLINK \l "_Toc138137979" T-COFFEE_LIB_FORMAT_01  PAGEREF _Toc138137979 \h 53  HYPERLINK \l "_Toc138137980" T-COFFEE_LIB_FORMAT_02  PAGEREF _Toc138137980 \h 53  HYPERLINK \l "_Toc138137981" Library List  PAGEREF _Toc138137981 \h 54  HYPERLINK \l "_Toc138137982" Substitution matrices.  PAGEREF _Toc138137982 \h 54  HYPERLINK \l "_Toc138137983" ClustalW Style [Deprecated]  PAGEREF _Toc138137983 \h 54  HYPERLINK \l "_Toc138137984" BLAST Format [Recommended]  PAGEREF _Toc138137984 \h 54  HYPERLINK \l "_Toc138137985" Sequences Weights  PAGEREF _Toc138137985 \h 54  HYPERLINK \l "_Toc138137986" Known Problems  PAGEREF _Toc138137986 \h 56  HYPERLINK \l "_Toc138137987" Technical Notes  PAGEREF _Toc138137987 \h 57  HYPERLINK \l "_Toc138137988" Development  PAGEREF _Toc138137988 \h 57  HYPERLINK \l "_Toc138137989" Command Line List  PAGEREF _Toc138137989 \h 57  HYPERLINK \l "_Toc138137990" To Do  PAGEREF _Toc138137990 \h 59  License and Terms of Use T-Coffee is distributed under the Gnu Public License 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 is no restriction to its use, either in an academic or a non academic environment. T-Coffee code can be re-used freely Our philosophy is that code is meant to be re-used, including ours. No permission is needed, although we are always happy to receive pieces of improved code. T-Coffee can be incorporated in any pipeline: Plug-in/Plug-out 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. Addresses and Contacts Contributors T-coffee is developed, maintained, monitored, used and debugged by a dedicated team that include: Cdric Notredame Fabrice Armougom Des Higgins Sebastien Moretti Orla OSullivan Eamon OToole Olivier Poirot Karsten Suhre Vladimir Keduas Iain Wallace Andreas Wilm Addresses We are always very eager to get some user feedback. Please do not hesitate to drop us a line at:  HYPERLINK "mailto:cedric.notredame@europe.com" cedric.notredame@europe.com 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 Citations 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").  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=10964570" Notredame C, Higgins DG, Heringa J. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=10964570" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu10964570);" \t "_self" Links T-Coffee: A novel method for fast and accurate multiple sequence alignment. J Mol Biol. 2000 Sep 8;302(1):205-17. PMID: 10964570 [PubMed - indexed for MEDLINE]Other useful publications include: T-Coffee  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=15215460" Claude JB, Suhre K, Notredame C, Claverie JM, Abergel C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=15215460" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu15215460);" \t "_self" Links CaspR: a web server for automated molecular replacement using homology modelling. Nucleic Acids Res. 2004 Jul 1;32(Web Server issue):W606-9. PMID: 15215460 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=15215345" Poirot O, Suhre K, Abergel C, O'Toole E, Notredame C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=15215345" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu15215345);" \t "_self" Links 3DCoffee@igs: a web server for combining sequences and structures into a multiple sequence alignment. Nucleic Acids Res. 2004 Jul 1;32(Web Server issue):W37-40. PMID: 15215345 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=15201059" O'Sullivan O, Suhre K, Abergel C, Higgins DG, Notredame C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=15201059" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu15201059);" \t "_self" Links 3DCoffee: combining protein sequences and structures within multiple sequence alignments. J Mol Biol. 2004 Jul 2;340(2):385-95. PMID: 15201059 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=12824354" Poirot O, O'Toole E, Notredame C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=12824354" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu12824354);" \t "_self" Links Tcoffee@igs: A web server for computing, evaluating and combining multiple sequence alignments. Nucleic Acids Res. 2003 Jul 1;31(13):3503-6. PMID: 12824354 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=11301309" Notredame C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=11301309" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu11301309);" \t "_self" Links Mocca: semi-automatic method for domain hunting. Bioinformatics. 2001 Apr;17(4):373-4. PMID: 11301309 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=10964570" Notredame C, Higgins DG, Heringa J. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=10964570" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu10964570);" \t "_self" Links T-Coffee: A novel method for fast and accurate multiple sequence alignment. J Mol Biol. 2000 Sep 8;302(1):205-17. PMID: 10964570 [PubMed - indexed for MEDLINE]  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=9682054" Notredame C, Holm L, Higgins DG. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=9682054" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu9682054);" \t "_self" Links COFFEE: an objective function for multiple sequence alignments. Bioinformatics. 1998 Jun;14(5):407-22. PMID: 9682054 [PubMed - indexed for MEDLINE] Mocca  HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=11301309" Notredame C. HYPERLINK "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Display&dopt=pubmed_pubmed&from_uid=11301309&tool=ExternalSearch" Related Articles,  HYPERLINK "javascript:PopUpMenu2_Set(Menu11301309);" \t "_self" Links Mocca: semi-automatic method for domain hunting. Bioinformatics. 2001 Apr;17(4):373-4. PMID: 11301309 [PubMed - indexed for MEDLINE]CORE  HYPERLINK "http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf" http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf Other Contributions 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 rationnaly Bug Reports and Feedback -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 Standard Installation of T-Coffee 1-decompress distribution.tar.gz gunzip distribution.tar.gz 2-untar distribution.tar tar -xvf distribution.tar 3-This will create the distribution directory with the following structure: distribution/bin distribution/doc/t_coffee_doc.pdf,t_coffee_doc.html distribution/t_coffee_source distribution/example distribution/html 4-go into the main directory and type: ./install You will know the installation proceeded completely with the mention: Installation of t_coffee Successful 5-When this is done, the t_coffee executable appears as: bin/t_coffee You can copy this file to the location where you store your executables (often something like (~/bin/). Alternatively, you can also add the current location to your path (Not recommended) using the following: set path = ($path .
) Note: The latest t_coffee distribution (2.15 and higher) is self contained and only requires one executable. You may still require external modules (sap, blast, ClustalW) if you wish to use another mode than the default. Note: When updating, make sure to remove the old distribution and any associated program from your path. 6-If you have PDB installed: Assuming you have a standard PDB installation in your file system setenv (or export) PDB_DIR /data/structures/all/pdb/ OR setenv (or export) PDB_DIR /structures/divided/pdb/ If you do not bhave 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. Installation of M-Coffee M-Coffee is a special mode of T-Coffee that makes it possible to combine the output of many multiple sequence alignment packages. 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/ ---------------------------------------------------------- MAFFT  HYPERLINK "http://www.biophys.kyoto-" http://www.biophys.kyoto-u.ac.jp/~katoh/programs/align/mafft/ ---------------------------------------------------------- Dialign-T  HYPERLINK "http://dialign-t.gobics.de/" http://dialign-t.gobics.de/ ---------------------------------------------------------- PCMA  HYPERLINK "ftp://iole.swmed.edu/pub/PCMA/" ftp://iole.swmed.edu/pub/PCMA/ 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 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 For two 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 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 Installation of APDB and iRMSD 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 Installation of seq_reformat 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 Installation of extract_from_pdb 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 apdb h Extract_from_pdb requires wget in order to automatically fetch PDB structures. Installation of 3D-Coffee In order to make the most out of T-Coffee, you will need to install the following packages: Package Function =================================================== --------------------------------------------------- wget 3DCoffee Automatic Downloading of Structures Remote use of the Fugue server --------------------------------------------------- sap structure/structure comparisons (obtain it from W. Taylor, NIMR-MRC). --------------------------------------------------- Blast  HYPERLINK "http://www.ncbi.nih.nlm.gov" www.ncbi.nih.nlm.gov --------------------------------------------------- Fugue protein to structure alignment program http://www-cryst.bioc.cam.ac.uk/fugue/download.html Once the package is installed, make sure make sure that the executable is on your path, so that t_coffee can find it automatically. Installing Fugue for T-Coffee Uses a standard fugue installation and install the follwing packages: joy, melody, fugueali, sstruc, hbond Copy some data into: cp fugue/classdef.dat /data/fugue/SUBST/classdef.dat OR Setenv MELODY_CLASSDEF= Setenv MELODY_SUBST=fugue/allmat.dat All the other configuration files must be in the right location. Installation of R-Coffee 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/ --------------------------------------------------- M-Coffee T-Coffee and the most common MSA Packages (cf M-Coffee in this installation guide) Installing Consan for R-Coffee 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. T-COFFEE 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 special_mode=dna is not needed or supported anymore PROMPT: t_coffee sample_dnaseq1.fasta type=dna M-Coffee 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 environement 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 special_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. R-Coffee 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: PROMPT: 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 special_mode mrcoffee If you want to decide yourself which methods should be combined by R-Coffee, run: PROMPT: t_coffee sample_rnaseq1.fasta special_mode rcoffee -method lalign_id_pair slow_pair iRMSD and APDB 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 MOCCA 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 (.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. -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: -special_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 thats 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 Environment Variables It is possible to modify T-Coffees behavior by setting any of the following environement variables. On the bash shell, use export VAR=value. On the cshell, use set $VAR=xxx DIR_4_TCOFFEE 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. TMP_4_TCOFFEE By default this variable is set to $HOME/.t_coffee/tmp. This is where T-Coffee stores temporary files. CACHE_4_TCOFFEE 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.... NO_ERROR_REPORT_4_TCOFFEE 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). PDB_DIR Indicate the location of your local PDB installation. NO_WARNING_4_TCOFFEE Suppresses all the warnings. Well Behaved Parameters Separation You can use any kind of separator you want (i.e. ,; =). 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. Posix T-Coffee is not POSIX compliant. Entering the right parameters 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 > -special_mode This means that -parameters supersede all the others, while parameters provided via -special mode are the weakest. Parameters Syntax No Flag If no flag is used 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. -parameters 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 -t_coffee_defaults Usage: -t_coffee_defaults= 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. 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 -special_mode Usage: -special_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 -score [Deprecated] Usage: -score Default: not used Toggles on the evaluate mode and causes t_coffee to evaluates a precomputed alignment provided via -infile=. 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 -special_mode=evaluate. -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=. 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: special_mode=evaluate. PROMPT: t_coffee infile=sample_aln1.aln -special_mode=evaluate PROMPT: t_coffee infile=sample_seq1.aln in Lsample_lib1.tc_lib special_mode=evaluate -convert [cw] 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. -do_align [cw] Usage: -do_align Default: turned on Special Parameters -version Usage: -version Default: not used Returns the current version number -check_configuration Usage: -check_configuration Default: not used Checks your system to determine whether all the programs T-Coffee can interact with are installed. -cache Usage: -cache=> 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. -update Usage: -update Default: turned off Causes a wget access that checks whether the t_coffee version you are using needs updating. -full_log Usage: -full_log= Default: turned off Causes t_coffee to output a full log file that contains all the input/output files. -other_pg Usage: -other_pg= 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 Input Sequence Input -infile [cw] 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) -get_type Usage: -get_type Default: turned off Forces t_coffee to identify the sequences type (PROTEIN, DNA). -type [cw] Usage: -type=DNA PROTEIN DNA_PROTEIN Default: -type= This flag sets the type of the sequences. If omitted, the type is guessed automatically. This flag is compatible with ClustalW. Warning: In case of low complexity or short sequences, it is recommended to set the type manually. -seq Usage: -seq=[,] Default: none -seq is now the recommended flag to provide your sequences. It behaves mostly like the -in flag. -seq_source Usage: -seq_source= 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). Structure Input -pdb Usage: -pdb=,[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 Tree Input -usetree Usage: -usetree= 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. Structures, Sequences Methods and Library Input via the in Flag 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) -lib Libraries (L) -in Usage: -in=[,] 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. Profile Input -profile Usage: -profile=[,] 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) -profile1 [cw] Usage: -profile1=[], one name only Default: no default Similar to the previous one and was provided for compatibility with ClustalW. -profile2 [cw] Usage: -profile1=[], one name only Default: no default Similar to the previous one and was provided for compatibility with ClustalW. Alignment Computation Library Computation: Methods -lalign_n_top Usage: -lalign_n_top= Default: -lalign_n_top=10 Number of alignment reported by the local method (lalign). -align_pdb_param_file Unsuported -align_pdb_hasch_mode Unsuported Library Computation: Extension -lib_list [Unsupported] Usage: -lib_list= 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). -do_normalise 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. -extend 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. -extend_mode Usage: -extend= 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 -max_n_pair Usage: -max_n_pair= 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). -seq_name_for_quadruplet Usage: Unsupported -compact Usage: Unsupported -clean Usage: Unsupported -maximise Usage: Unsupported -do_self 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. -seq_name_for_quadruplet Usage: Unsupported -weight Usage: -weight= or Default: -weight=sim Weight defines the way alignments are weighted when turned into a library. Overweighting can be obtained with the OW 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 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 Tree Computation -distance_matrix_mode Usage: -distance_matrix_mode= 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 -quicktree [CW] 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 Pair-wise Alignment Computation 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 -dp_mode Usage: -dp_mode= 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. -ktuple Usage: -ktuple= 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.. -ndiag Usage: -ndiag= 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. -diag_mode Usage: -diag_mode= 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). -diag_threshold Usage: -diag_threshold= 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). -sim_matrix Usage: -sim_matrix= 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. -matrix [CW] Usage: -matrix= 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. -nomatch Usage: -nomatch= 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). -gapopen Usage: -gapopen= 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. -gapext Usage: -gapext= Default: -gapext=0 Indicates the penalty applied for extending a gap (cf -gapopen) -fgapopen Unsupported -fgapext Unsupported -cosmetic_penalty Usage: -cosmetic_penalty= 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. -tg_mode 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. Weighting Schemes -seq_weight Usage: -seq_weight=> 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). Multiple Alignment Computation -msa_mode Usage: -msa_mode= Default: -evaluate_mode=tree Unsupported -one2all Usage: -one2all= 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= 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. -profile_mode Usage: -profile_mode= Default: -profile_mode=cw_profile_profile When profile_comparison=profile, this flag selects a profile scoring function. Alignment Post-Processing -clean_aln 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, -clean_threshold Usage: -clean_threshold=<0-9> Default:-clean_aln=1 See -clean_aln for details. -clean_iteration Usage: -clean_iteration= Default:-clean_iteration=1 See -clean_aln for details. -clean_evaluation_mode Usage: -clean_iteration= 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. -iterate Usage: -iterate= 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. CPU Control Multithreading -multi_thread [NOT Supported] Usage: -multi_thread= Default: 0 Specifies that the program should be used in multithreading mode. N specifies the number of processors available. PROMPT: t_coffee sample_seq2.fasta -multi_thread 4 If you are using a quadriprocessor Limits -mem_mode Usage: deprecated -ulimit Usage: -ulimit= 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. -maxlen Usage: -maxlen= Default: -maxlen=1000 Indicates the maximum length of the sequences. Aligning more than 100 sequences with DPA -maxnseq Usage: -maxnseq= Default: -maxnseq=50 Indicates the maximum number of sequences before triggering the use of t_coffee_dpa. -dpa_master_aln Usage: -dpa_master_aln= 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 -out -dpa_maxnseq Usage: -dpa_maxnseq= 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. -dpa_min_score1 Usage: -dpa_min_score1= 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. -dpa_min_score2 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. -dap_tree [NOT IMPLEMENTED] Usage: -dpa_tree= 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. Using Structures Generic -special_mode Usage: -special_mode=3dcoffee Default: turned off Runs t_coffee with the 3dcoffee mode (cf next section). -check_pdb_status 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. 3D Coffee: Using SAP 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 in=struc1.pdb,struc2.pdb,struc3.pdb,Msap_pair Will combine the pairwise alignments produced by SAP. There are currently two 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. 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 special_mode 3dcoffee seq 3d_sample3.fasta -template_file template_file.template Using/finding PDB templates for the Sequences -template_file Usage: -template_file = 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: 1-File name This file contains the sequence/template association it uses a FASTA-like format, as follows: > _P_ > _G_ > _R_ > _F_ 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_ 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= -outfile= 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= -outfile= 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 -struc_to_use Usage: -struc_to_use= Default: -struc_to_use=NULL Restricts the 3Dcoffee to a set of pre-defined structures. Multiple Local Alignments 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. -domain/-mocca 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 . 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. -start Usage: -start= 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. -len Usage: -len= Default: not set This flag indicates the length of the portion of sequence that will be used as a template. -scale Usage: -scale= 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 (Increase sensitivity ( 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: |:start Type S to save the current alignment, and extract a new motif. Type X when you are done. Output Control Generic 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 TYPE FORMAT -no_warning Usage: -no_warning Default: Switched off Suppresseswarning output. Alignments -outfile Usage: -outfile= Default:-outfile=default Indicates the name of the alignment output by t_coffee. If the default is used, the alignment is named .aln -output Usage: -output= Default:-output=clustalw Indicates the format used for outputting the -outfile. Supported formats are: clustalw_aln, clustalw : ClustalW format. gcg, msf_aln : MSF alignment. pir_aln : pir alignment. fasta_aln : fasta alignment. phylip : Phylip format. pir_seq : pir sequences (no gap). fasta_seq : fasta sequences (no gap). As well as: score_ascii : causes the output of a reliability flag score_html : causes the output to be a reliability plot in HTML score_pdf : idem in PDF (if ps2pdf is installed on your system). score_ps : idem in postscript. More than one format can be indicated: PROMPT: t_coffee sample_seq1.fasta -output=clustalw,gcg, score_html A publication describing the CORE index is available on:  HYPERLINK "http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf" http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf -outseqweight Usage: -outseqweight= Default: not used Indicates the name of the file in which the sequences weights should be saved.. -case Usage: -case= Default: -case=keep Instructs the program on the case to be used in the output file (Clustalw uses upper case). The default keeps the case and makes it possible to maintain a mixture of upper and lower case residues. If you need to change the case of your file, you can use seq_reformat: PROMPT: t_coffee other_pg seq_reformat in sample_aln1.aln action +lower output clustalw -cpu Usage: deprecated -outseqweight Usage: -outseqweight= Default: -outseqweight=no Will cause the program to output the weights associated with every sequence in the dataset. -outorder [cw] Usage: -outorder= Default:-outorder=input Sets the order of the sequences in the output alignment: -outorder=input means the sequences are kept in the original order. -outorder=aligned means the sequences come in the order indicated by the tree. This order can be seen as a one-dimensional projection of the tree distances. outdorder=Filename is a legal fasta file, whose order will be used in the final alignment. -inorder [cw] Usage: -inorder= Default:-inorder=aligned Multiple alignments based on dynamic programming depend slightly on the order in which the incoming sequences are provided. To prevent this effect sequences are arbitrarily sorted at the beginning of the program (-inorder=aligned). However, this affects the sequence order within the library. You can switch this off by ststing inorder=input. -seqnos Usage: -seqnos= Default:-seqnos=off Causes the output alignment to contain residue numbers at the end of each line: T-COFFEE seq1 aaa---aaaa--------aa 9 seq2 a-----aa-----------a 4 seq1 a-----------------a 11 seq2 aaaaaaaaaaaaaaaaaaa 19 Libraries Although, it does not necessarily do so explicitly, T-Coffee always end up combining libraries. Libraries are collections of pairs of residues. Given a set of libraries, T-Coffee makes an attempt to assemble the alignment with the highest level of consistence. You can think of the alignment as a timetable. Each library pair would be a request from students or teachers, and the job of T-Coffee would be to assemble the time table that makes as many people as possible happy -out_lib Usage: -out_lib= Default:-out_lib=default Sets the name of the library output. Default implies .tc_lib -lib_only Usage: -lib_only Default: unset Causes the program to stop once the library has been computed. Must be used in conjunction with the flag out_lib Trees -newtree Usage: -newtree= Default: No file specified Indicates the name of the file into which the guide tree will be written. The default will be .dnd, or . The tree is written in the parenthesis format known as newick or New Hampshire and used by Phylips (see the format section). Do NOT confuse this guide tree with a phylogenetic tree. Reliability Estimation CORE Computation The CORE is an index that indicates the consistency between the library of piarwise alignments and the final multiple alignment. Our experiment indicate that the higher this consistency, the more reliable the alignment. A publication describing the CORE index can be found on:  HYPERLINK "http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf" http://igs-server.cnrs-mrs.fr/~cnotred/Publications/Pdf/core.pp.pdf -evaluate_mode Usage: -evaluate_mode= Default: -evaluate_mode=t_coffee_fast This flag indicates the mode used to normalize the t_coffee score when computing the reliability score. t_coffee_fast: Normalization is made using the highest score in the MSA. This evaluation mode was validated and in our hands, pairs of residues with a score of 5 or higher have 90 % chances to be correctly aligned to one another. t_coffee_slow: Normalization is made using the library. This usually results in lower score and a scoring scheme more sensitive to the number of sequences in the dataset. Note that this scoring scheme is not any more slower, thanks to the implementation of a faster heuristic algorithm. t_coffee_non_extended: the score of each residue is the ratio between the sum of its non extended scores with the column and the sum of all its possible non extended scores. These modes will be useful when generating colored version of the output, with the output flag: PROMPT: t_coffee sample_seq1.fasta evaluate_mode t_coffee_slow output score_ascii, score_html PROMPT: t_coffee sample_seq1.fasta evaluate_mode t_coffee_fast output score_ascii, score_html PROMPT: t_coffee sample_seq1.fasta evaluate_mode t_coffee_non_extended output score_ascii, score_html Generic Output -run_name Usage: -run_name= Default: no default set This flag causes the prefix to be replaced by when renaming the default output files. -quiet Usage: -quiet=. Default:-quiet=stderr Redirects the standard output to either a file. -quiet on its own redirect the output to /dev/null. -align [CW] This flag indicates that the program must produce the alignment. It is here for compatibility with ClustalW. APDB/iRMSD Parameters Warning: These flags will only work within the APDB package that can be invoked via the other_pg parameter of T-Coffee: t_coffee other_pg apdb aln -quiet [Same as T-Coffee] -run_name [Same as T-Coffee] -aln Usage: -aln=. Default:none Indicates the name of the file containing the sequences that need to be evaluated. The sequences whose structure is meant to be used must be named according to their PDB identifier. The format can be FASTA, CLUSTAL or any of the formats supported by T-Coffee. APDB only evaluates residues in capital and ignores those in lower case. If your sequences are in lower case, you can upper case them using seq_reformat: PROMPT: t_coffee other_pg seq_reformat in 3d_sample4.aln action +upper output clustalw > 3d_sample4.cw_aln The alignment can then be evaluated using the defaultr of APDB: PROMPT: t_coffee other_pg apdb aln 3d_sample4.aln The alignment can contain as many structures as you wish. -n_excluded_nb Usage: -n_excluded_nb=. Default:1 When evaluating the local score of a pair of aligned residues, the residues immediately next to that column should not contribute to the measure. By default the first to the left and first to the right are excluded. -maximum_distance Usage: -maximum_distance=. Default:10 Size of the neighborhood considered around every residue. If .-local_mode is set to sphere, -maximum_distance is the radius of a sphere centered around each residue. If local_mode is set to window, then maximum_distance is the size of the half window (i.e. window_size=-maximum_distance*2+1). -similarity_threshold Usage: -similarity_threshold=. Default:70 Fraction of the neighborhood that must be supportive for a pair of residue to be considered correct in APDB. The neighborhood is a sphere defined by maximum_distance, and the support is defined by md_threshold. -local_mode Usage: -local_mode=. Default:sphere Defines the shape of a neighborhood, either as a sphere or as a window. -filter Usage: -filter=<0.00-1.00>. Default:1.00 Defines the centiles that should be kept when making the local measure. Foir instance, -filter=0.90 means that the the 10 last centiles will be removed from the evaluation. The filtration is carried out on the iRMSD values. -print_rapdb [Unsupported] Usage: -print_rapdb (FLAG) Default:off This causes the prints out of the exact neighborhood of every considered pair of residues. -outfile [Same as T-Coffee] This flag is meant to control the output name of the colored APDB output. This file will either display the local APDB score or the local iRMD, depending on the value of color_mode. The default format is defined by ouptut and is score_html. -color_mode Usage: -color_mode= Default:apdb This flag is meant to control the colored APDB output (local score). This file will either display the local APDB score or the local iRMD. Building a Server We maintain a T-Coffee server (www.tcoffee.org). We will be pleased to provide anyone who wants to set up a similar service with the sources Environment Variables T-Coffee stores a lots of information in locations that may be unsuitable when running a server. By default, T-Coffee will generate and rely on the follwing directory structure: /home/youraccount/ #HOME_4_TCOFFEE HOME_4_TCOFFEE/.t_coffee/ #DIR_4_TCOFFEE DIR_4_TCOFFEE/cache #CACHE_4_TCOFFEE DIR_4_TCOFFEE/tmp #TMP_4_TCOFFEE DIR_4_TCOFFEE/methods #METHOS_4_TCOFFEE DIR_4_TCOFFEE/mcoffee #MCOFFEE_4_TCOFFEE By default, all these directories are automatically created, following the dependencies suggested here. The first step is the determination of the HOME. By default the program tries to use HOME_4_TCOFFEE, then the HOME variable and TMP or TEMP if HOME is not set on your system or your account. It is your responsibility to make sure that one of these variables is set to some valid location where the T-Coffee process is allowed to read and write. If no valid location can be found for HOME_4_TCOFFEE, the program exits. If you are running T-Coffee on a server, we recommend to hard set the following locations, where your scratch is a valid location. HOME_4_TCOFFEE=your scratch TMP_4_TCOFFEE=your scratch DIR_4_TCOFFEE=your scratch CACHE_4_TCOFFEE=your scratch NO_ERROR_REPORT_4_TCOFFEE=1 Note that it is a good idea to have a cron job that cleans up this scratch area, once in a while. Output of the .dnd file. A common source of error when running a server: T-Coffee MUST output the .dnd file because it re-reads it to carry out the progressive alignment. By default T-Coffee outputs this file in the directory where the process is running. If the T-Coffee process does not have permission to write in that directory, the computation will abort... To avoid this, simply specify the name of the output tree: -newtree= Chose the name so that two processes may not over-write each other dnd file. Permissions The t_coffee process MUST be allowed to write in some scratch area, even when it is ran by Mr nobody... Make sure the /tmp/ partition is not protected. Other Programs T-Coffee may call various programs while it runs (lalign2list by defaults). Make sure your process knows where to find these executables. Formats Parameter files Parameter files used with -parameters, -t_coffee_defaults, -dali_defaults... Must contain a valid parameter string where line breaks are allowed. These files cannot contain any comment, the recommended format is one parameter per line: =,.... =..... Sequence Name Handling Sequence name handling is meant to be fully consistent with ClustalW (Version 1.75). This implies that in some cases the names of your sequences may be edited when coming out of the program. Five rules apply: Naming Your Sequences the Right Way 1-No Space Names that do contain spaces, for instance: >seq1 human_myc will be turned into >seq1 It is your responsibility to make sure that the names you provide are not ambiguous after such an editing. This editing is consistent with Clustalw (Version 1.75) 2-No Strange Character Some non alphabetical characters are replaced with underscores. These are: ';:()' Other characters are legal and will be kept unchanged. This editing is meant to keep in line with Clustalw (Version 1.75). 3-> is NEVER legal (except as a header token in a FASTA file) 4-Name length must be below 100 characters, although 15 is recommended for compatibility with other programs. 5-Duplicated sequences will be renamed (i.e. sequences with the same name in the same dataset) are allowed but will be renamed according to their original order. When sequences come from multiple sources via the in flag, consistency of the renaming is not guaranteed. You should avoid duplicated sequences as they will cause your input to differ from your output thus making it difficult to track data. Automatic Format Recognition Most common formats are automatically recognized by t_coffee. See -in and the next section for more details. If your format is not recognized, use readseq or clustalw to switch to another format. We recommend Fasta. Structures PDB format is recognized by T-Coffee. T-Coffee uses extract_from_pdb (cf other_pg flag). extract_from_pdb is a small embeded module that can be used on its own to extract information from pdb files. RNA Structures RNA structures are expressed as T-Coffee libraries, with each line indicating two paired residues. Sequences Sequences can come in the following formats: fasta, pir, swiss-prot, clustal aln, msf aln and t_coffee aln. These formats are the one automatically recognized. Please replace the '*' sign sometimes used for stop codons with an X. Alignments Alignments can come in the following formats: msf, ClustalW, Fasta, Pir and t_coffee. The t_coffee format is very similar to the ClustalW format, but slightly more flexible. Any interleaved format with sequence name on each line will be correctly parsed: [Facultative]n [Required] [Facultative]n [Required] [Facultative]n [Required] [Facultative]n [Required] [Facultative]n An empty line is a line that does NOT contain amino-acid. A line that contains the ClustalW annotation (.:*) is empty. Spaces are forbidden in the name. When the alignment is being read, non character signs are ignored in the sequence field (such as numbers, annotation). Note: a different number of lines in the different blocks will cause the program to crash or hang. Libraries T-COFFEE_LIB_FORMAT_01 This is currently the only supported format. ! TC_LIB_FORMAT_01 !Comment (!Comment)n #Si1 Si2 Ri1 Ri2 V1 (V2, V3) #1 2 12 13 99 (12/0 vs 13/1, weight 99) 12 14 70 15 16 56 #1 3 12 13 99 12 14 70 15 16 56 !SEQ_1_TO_N Si1: index of Sequence 1 Ri1: index of residue 1 in seq1 V1: Integer Value: Weight V2, V3: optional values Note 1: There is a space between the ! And SEQ_1_TO_N Note 2: The last line (! SEQ_1_TO_N) indicates that: Sequences and residues are numbered from 1 to N, unless the token SEQ_1_TO_N is omitted, in which case the sequences are numbered from 0 to N-1, and residues are from 1 to N. Residues do not need to be sorted, and neither do the sequences. The same pair can appear several times in the library. For instance, the following file would be legal: #1 2 12 13 99 #1 2 15 16 99 #1 1 12 14 70 It is also poosible to declare ranges of resdues rather than single pairs. For instance, the following: #0 1 +BLOCK+ 10 12 14 99 +BLOCK+ 15 30 40 99 #0 2 15 16 99 #0 1 12 14 70 The first statement BLOCK declares a BLOCK of length 10, that starts on position 12 of sequence 1 and position 14 of sequence 2 and where each pair of residues within the block has a score of 99. The second BLOCK starts on residue 30 of 1, residue 40 of 2 and extends for 15 residues. Blocks can overalp and be incompatible with one another, just like single constraints. T-COFFEE_LIB_FORMAT_02 A simpler format is being developed, however it is not yet fully supported and is only mentioned here for development purpose. ! TC_LIB_FORMAT_02 #S1 SEQ1 [OPTIONAL] #S2 SEQ2 [OPTIONAL] ... !comment [OPTIONAL] S1 R1 Ri1 S2 R2 Ri2 V1 (V2 V3) => N R1 Ri1 S2 R2 Ri2 V1 (V2 V3) ... S1, S2: name of sequence 1 and 2 SEQ1: sequence of S1 Ri1, Ri2: index of the residues in their respective sequence R1, R2: Residue type V1, V2, V3: integer Values (V2 and V3 are optional) Value1, Value 2 and Value3 are optional. Library List These are lists of pairs of sequences that must be used to compute a library. The format is: 2 hamg2 globav 3 hamgw hemog singa ... Substitution matrices. If the required substitution matrix is not available, write your own in a file using the following format: ClustalW Style [Deprecated] # CLUSTALW_MATRIX FORMAT $ v1 v2 v3 v4 v5 v6 ... $ v1, v2... are integers, possibly negatives. The order of the amino acids is: ABCDEFGHIKLMNQRSTVWXYZ, which means that v1 is the substitution value for A vs A, v2 for A vs B, v3 for B vs B, v4 for A vs C and so on. BLAST Format [Recommended] # BLAST_MATRIX FORMAT # ALPHABET=AGCT A G C T A 0 1 2 3 G 0 2 3 4 C 1 1 2 3 ... The alphabet can be freely defined Sequences Weights Create your own weight file, using the -seq_weight flag: # SINGLE_SEQ_WEIGHT_FORMAT_01 seq_name1 v1 seq_name2 v2 ... No duplicate allowed. Sequences not included in the set of sequences provided to t_coffee will be ignored. Order is free. V1 is a float. Un-weighted sequences will see their weight set to 1. Known Problems 1-Sensitivity to sequence order: It is difficult to implement a MSA algorithm totally insensitive to the order of input of the sequences. In t_coffee, robustness is increased by sorting the sequences alphabetically before aligning them. Beware that this can result in confusing output where sequences with similar name are unexpectedly close to one another in the final alignment. 2-Nucleotides sequences with long stretches of Ns will cause problems to lalign, especially when using Mocca. To avoid any problem, filter out these nucleotides before running mocca. 3-Stop codons are sometimes coded with '*' in protein sequences. This will cause the program to crash or hang. Please replace the '*' signs with an X. 4-Results can differ from one architecture to another, due rounding differences. This is caused by the tree estimation procedcure. If you want to make sure an alignment is reproducible, you should keep the associated dendrogram. Technical Notes These notes are only meant for internal development. Development The following examples are only meant for internal development, and are used to insure stability from release to release profile2list prf1: profile containing one structure prf2: profile containing one structure PROMPT: t_coffee Rsample_profile1.aln,Rsample_profile2.aln -special_mode=3dcoffee -outfile=aligned_prf.aln Command Line List These command lines have been checked before every release (along with the other CL in this documentation: -external methods; PROMPT: t_coffee sample_seq1.fasta in=Mclustalw_pair,Mclustalw_msa,Mslow_pair outfile=clustal_text -fugue_client PROMPT: t_coffee in Ssample_seq5.fasta Pstruc4.pdb Mfugue_pair -A list of command lines kindly provided by James Watson (used to crash the pg before version 3.40) PROMPT: t_coffee -in Sseq.fas P2PTC Mfugue_pair PROMPT: t_coffee -in S2seqs.fas Mfugue_pair -template_file SELF_P_ PROMPT: t_coffee -special_mode 3dcoffee -in Sseq.fas P2PTC PROMPT: t_coffee -special_mode 3dcoffee -in S2seqs.fas -template_file SELF_P_ -A list of command lines that crashed the program before 3.81 PROMPT: t_coffee sample_seq6.fasta in Mfast_pair Msap_pair Mfugue_pair template_file template_file6.template -A command line to read relaxed pdb files... PROMPT: t_coffee in Msap_pair Ssample_seq7.fasta template_file template_file7.template weight 1001 out_lib test_lib7.tc_lib lib_only -Parsing of MARNA libraries PROMPT: t_coffee in Lmarna.tc_lib outfile maran.test -Parsing of long sequence lines: PROMPT: t_coffee in Asample_aln5.aln outfile test.aln To Do -implement UPGMA tree computation -implement seq2dpa_tree -debug dpa -Reconciliate sequences and template when reading the template -Add the server command lines to the checking procedure      PAGE 1 T-Coffee reference manual  PAGE 4 Design Customization PAGE 15   eu  / 0 7 Z r z { | } ٲ٨p]$jhoLh0JUmHnHu"hoLh|6CJOJQJ]aJ+jhoLh|6CJOJQJU]aJhoLh|@ hoLh e jhoLh|h8 hoLh 2huh+hoLh|@CJKH hoLh hoLh}Z& hoLh)yMh?4h?QCJaJ hoLh| hoLhN  e/ { | *  1 }  \ W %  % Ygd|5gd8gd|\gd)yM ]]^gd|\gd|Agd?QVgd|ZƤ  $ % & ' ( ) * + , H I J K  ˸񪘪eK񪘪2jhoLh>*B*UmHnHphu=hoLh5B*CJOJPJQJ\]mHnHphtHu&j}hoLhUmHnHu#jhoLhUmHnHuhoLhmHnHu$jhoLh0JUmHnHu2jhoLh>*B*UmHnHphuhoLhmHnHuhoLh0JmHnHu         % & 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