Universität Bielefeld - Technische Fakultät - AG Praktische Informatik - FSPM - Strukturbildungsprozesse
GSF Research Center - GSF, AG BioDV - GSF, Institute of Biomathematics and Biometry

DIALIGN 2

User Guide

Developed by:

Burkhard Morgenstern, Thomas Werner, Klaus Hahn
GSF - National Research Center for Environment and Health,
Ingolstädter Landstr. 1, 85764 Neuherberg, Germany
Andreas Dress
Fakultät für Mathematik, Universität Bielefeld,
33501 Bielefeld, Postfach 100131, Germany
Kornelie Frech
Genomatix Software GmbH,
Karlstr. 55, 80333 München, Germany

at GSF, AG BioDV, University of Bielefeld - FSPM, GSF, Institute of Biomathematics and Biometry and North Carolina State University, Department of Genetics

E-mail contact: morgenstern@gsf.de

Important Note:

Use of DIALIGN 2 is subject to the copyright notice (COPYRIGHT).

Distribution of copies of this user guide to all users of the program is explicitly encouraged since it will facilitate using DIALIGN.

References:

The basic ideas behind DIALIGN are described in

B. Morgenstern, A. Dress, T. Werner (1996).
Multiple DNA and protein sequence alignment based on segment-to-segment comparison.
Proc. Natl. Acad. Sci. USA 93, pp. 12098 - 12103.

B. Morgenstern, K. Frech, A. Dress, T. Werner (1998).
DIALIGN: Finding local similarities by multiple sequence alignment.
Bioinformatics 14, in press.

The main improvement of DIALIGN 2 - compared to the first version of the program - is described in

B. Morgenstern, W.R. Atchley, K. Hahn, A. Dress (1998).
Segment-based scores for pairwise and multiple sequence alignments.
Proceedings of the Sixth International Conference on Intelligent Systems
for Molecular Biology (ISMB 98), in press.

Program Input:

There are two ways to run DIALIGN on your computer: You can run the program interactively or you can enter parameters via command line. In either case, sequences must be contained in a single

Sequence file:

DIALIGN requires an ASCII file containing the sequences to be aligned. Four different file formats are supported: IG, FASTA, EMBL and GCG-RSF format. The following is an example of the FASTA sequence file format:



        >HTL2  
        LDTAPCLFSDGSPQKAAYVLWDQTILQQDITPLPSHETHSAQKGELLALICGLRAAKPWP
        SLNIFLDSKYLIKYLHSLAIGAFLGTSAHQTLQAALPPLLQGKTIYLHHVRSHTNLPDPI
        STFNEYTDSLILAPL
        >MMLV   
        PDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAKALDAGTSAQRAELIALTQALKMAE
        GKKLNVYTDSRYAFATAHIHGEIYRRRGLLTSEGKEIKNKDEILALLKALFLPKRLSIIH
        CPGHQKGHSAEARGNRMADQAARKAAITETPDTSTLL
        >HEPB 
        RPGLCQVFADATPTGWGLVMGHQRMRGTFSAPLPIHTAELLAACFARSRSGANIIGTDNS
        VVLSRKYTSFPWLLGCAANWILRGTSFVYVPSALNPADDPSRGRLGLSRPLLRLPFRPTT
        GRTSLYADSPSVPSHLPDRVH
        >ECOL   
        MLKQVEIFTDGSCLGNPGPGGYGAILRYRGREKTFSAGYTRTTNNRMELMAAIVALEALK
        EHCEVILSTDSQYVRQGITQWIHNWKKRGWKTADKKPVKNVDLWQRLDAALGQHQIKWEW
        VKGHAGHPENERCDELARAAAMNPTLEDTGYQVEV

The first line for each sequence starts with ">" and contains the name of the sequence.

Options:

Sequence Type:
The user can decide if nucleic acid or protein sequences are to be aligned.
Threshold T:
As described in our papers, the program DIALIGN constructs alignments from gapfree pairs of similar segments of the sequences. Such segment pairs are referred to as `diagonals'.
Every possible diagonal is given a so-called weight reflecting the degree of similarity among the two segments involved. The overall score of an alignment is then defined as the sum of weights of the diagonals it consists of and the program tries to find an alignment with maximum score -- in other words: the program tries to find a consistent collection of diagonals with maximum sum of weights. This novel scoring scheme for alignments is the basic difference between DIALIGN and other global or local alignment methods. Note that DIALIGN does not employ any kind of gap penalty.
It is possible to use a threshold T for the quality of the diagonals. In this case, a diagonal is considered for alignment only if its `weight' exceeds this threshold. Regions of lower similarity are not aligned.
In the first version of the program (DIALIGN 1), this threshold was in many situations absolutely necessary to obtain meaningful alignments. By contrast, DIALIGN 2 should produce reasonable alignments without a threshold, i.e. with T = 0. This is the most important difference between DIALIGN 2 and the first version of the program.
Nevertheless, it is still possible to use a threshold T, so it is up to the user to experiment with this option.
Translation of `nucleotide diagonals' into `peptide diagonals':
If (possibly) coding nucleic acid sequences are to be aligned, DIALIGN optionally translates the compared `nucleic acid segments' to `peptide segments' according to the genetic code -- without (necessarily) presupposing any of the three possible reading frames, so all three of them get checked for significant similarity. In this case, the similarity among segments will be assessed on the `peptide level' rather than on the `nucleic acid level'.
We strongly recommend this option if nucleic acid sequences are expected to contain protein coding regions, as it will significantly increase the sensitivity of the alignment procedure in such cases.
`*' characters:
The user can specify the maximum number of `*' characters per column indicating the degree of local similarity among sequences in the DIALIGN alignment.

Entering parameters via command line:

If you want to enter options via command line, the program call is as follows:

 
  dialign [ -fn "out_file" ; -pln x ; -n ; -nt ; -sto ; -thr x ] "seq_file"

Here, "seq_file" is the name of the input sequence file. The meaning of the (optional) parameters in brackets is as follows:

`-fn "out_file"`	output file is named `"out_file"`
`-pln x`	maximum number of `*' characters representing
	degree of local similarity among sequences = `x`
`-n`	input sequences are nucleic acid sequences. No translation of diagonals.
`-nt`	input sequences are nucleic acid sequences and
	`nucleic acid diagonals' are translated into `peptide diagonals'.
`-sto`	Results are written to the standard output.
`-thr x`	Threshold T = x.

Similarity Matrix:

DIALIGN 2 uses the BLOSUM62 amino acid substitution matrix. In the current version, it is not possible to replace BLOSUM62 by other similarity matrices, since the probability values contained in the files n_prob and p_prob refer to the BLOSUM62 matrix.

Program Output:

DIALIGN creates a file containing

An alignment of the input sequences in DIALIGN format.
The same alignment in FASTA format.
A sequence tree in PHYLIP format. This tree is constructed by applying the UPGMA clustering method to the DIALIGN similarity scores. It roughly reflects the different degrees of similarity among sequences. For detailed phylogenetic analysis, we recommend the usual methods for phylogenetic reconstruction.

This is DIALIGN alignment format:

  
HTL2          1   ldtapcLFSD GS------PQ KAAYVLWDQT IL---QQDIT PLPSHethSA
MMLV          1   pdadhtwYTD GSSLLQEGQR KAGAAVTTET eviwaKALDA G---T---SA
HEPB          1   rpglcQVFAD AT------PT GWGLVMGHQR MR---GTFSA PLPIHt----
ECOL          1   mlkqvEIFTD GSCLGNPGPG GYGAILRYRG RE---KTFSA GytrT---TN
                                                                
                       ***** ********** ********** **   ***** *****   **
                        **** **      ** ********** **   ***** *****   **
                         *** **      ** ********** **   *****           
                                     ** ******                          
                                                                        


HTL2         42   QKGELLALIC GLRAAKPWPS LNIFLDSKYL IKYLHslaig aflgtsah--
MMLV         45   QRAELIALTQ ALKMAEgkk- LNVYTDSRYA FATAHIHGEI YRRRGLLTSE
HEPB         38   --AELLAACF Arsrsgan-- -IIGTDN--- ---------- ----------
ECOL         45   NRMELMAAIV ALEALKEHCE VILSTDSQYV RQGITQWIHN WKKRGWKTAD
                                                                
                  ********** ********** ********** ********** **********
                  ********** ********** ********** ********** **********
                     ******* ******     ********** *****                
                     ******* ******     ********** *****                
                                          ********                      


HTL2         90   -------QT- --LQAALPPL LQGKTIYLHH VRSHT----- -NLPDPISTF
MMLV         94   GKEIKNKDE- --ILALLKAL FLPKRLSIIH CPGHQ----- -KGHSAEARG
HEPB         60   ---------- ---SVVLSR- ---------- ---KYTSFPW LLGCAANWI-
ECOL         95   KKPVKNVDlw qrLDAALGQ- ---------- ---HQIKWEW VKGHAGHPE-
                                                                
                  *********    ******** ********** ********** **********
                  ********                                              
                         *                                              
                                                                        
                                                        


HTL2        124   NEYTDSLILA pl-------- ---------- ---------- ----------
MMLV        135   NRMADQAARK AAITETPDTS tll------- ---------- ----------
HEPB         82   LRGTSFVYVP SALNPADDPS rgrlglsrpl lrlpfrpttg rtslyadsps
ECOL        130   NERCDELARA AAMNPTledt gyqvev---- ---------- ----------
                                                                
                  ********** **********                                 
                  ********** ******                                     
                                                                        
                                                                        
                                                                        


HTL2        136   ----------
MMLV              ----------
HEPB        132   vpshlpdrvh
ECOL        156   ----------

Names of aligned sequences are shown on the left hand side of the alignment.

Numbers on the left hand side of the alignment denote the position of the first residue in a line within the respective sequence.

Capital letters denote aligned residues, i.e. residues involved in at least one of the `diagonals' the alignment consists of. Lower-case letters denote residues not belonging to any of these selected `diagonals'. They are not considered to be aligned by DIALIGN. Thus, if a lower-case letter is standing in the same column with other letters, this is pure chance; these residues are not considered to be homologous.
The number of `*' characters below the alignment reflects the degree of local similarity among sequences. More precisely: They represent the sum of `weights' of diagonals connecting residues at the respective position. The number of `*' characters is normalized such that regions of maximum similarity have N `*' characters per column. N can be specified by the user. By default, N = 5. Note that the number of `*' characters depicts the relative degree of similarity within an alignment, since in every alignment, the region of maximum similarity gets N `*' characters.

This is FASTA alignment format:


>HTL2
ldtapcLFSDGS------PQKAAYVLWDQTIL---QQDITPLPSHethSA
QKGELLALICGLRAAKPWPSLNIFLDSKYLIKYLHslaigaflgtsah--
-------QT---LQAALPPLLQGKTIYLHHVRSHT------NLPDPISTF
NEYTDSLILApl--------------------------------------
----------
>MMLV
pdadhtwYTDGSSLLQEGQRKAGAAVTTETeviwaKALDAG---T---SA
QRAELIALTQALKMAEgkk-LNVYTDSRYAFATAHIHGEIYRRRGLLTSE
GKEIKNKDE---ILALLKALFLPKRLSIIHCPGHQ------KGHSAEARG
NRMADQAARKAAITETPDTStll---------------------------
----------
>HEPB
rpglcQVFADAT------PTGWGLVMGHQRMR---GTFSAPLPIHt----
--AELLAACFArsrsgan---IIGTDN-----------------------
-------------SVVLSR--------------KYTSFPWLLGCAANWI-
LRGTSFVYVPSALNPADDPSrgrlglsrpllrlpfrpttgrtslyadsps
vpshlpdrvh
>ECOL
mlkqvEIFTDGSCLGNPGPGGYGAILRYRGRE---KTFSAGytrT---TN
NRMELMAAIVALEALKEHCEVILSTDSQYVRQGITQWIHNWKKRGWKTAD
KKPVKNVDlwqrLDAALGQ--------------HQIKWEWVKGHAGHPE-
NERCDELARAAAMNPTledtgyqvev------------------------
----------

This is PHYLIP tree format:

 
((HTL2:0.111024,
(MMLV:0.078471,
ECOL:0.078471):0.032554):0.121218,
HEPB:0.232242);

Trees can be visualized using the treetool program contained in the PHYLIP software package.

burkhard morgenstern (last modified: May 11, 1998)