#!/usr/bin/perl
# PROGRAM : clustalw.pl
# PURPOSE : Demonstrate possible uses of Bio::Tools::Run::Alignment::Clustalw.pm
# AUTHOR : Peter Schattner schattner@alum.mit.edu
# CREATED : Oct 06 2000
#
# INSTALLATION
#
# You will need to have installed clustalw and to ensure that Clustalw.pm can find it.
# This can be done in different ways (bash syntax):
# export PATH=$PATH:/home/peter/clustalw1.8
# or
# define an environmental variable CLUSTALDIR:
# export CLUSTALDIR=/home/peter/clustalw1.8
# or
# include a definition of an environmental variable CLUSTALDIR in every
# script that will use Clustal.pm.
# BEGIN {$ENV{CLUSTALDIR} = '/home/peter/clustalw1.8/'; }
#
# We are going to demonstrate 3 possible applications of Clustalw.pm:
# 1. Test effect of varying clustalw alignment parameter(s) on resulting alignment
# 2. Test effect of changing the order that sequences are added to the alignment
# on the resulting alignment
# 3. Test effect of incorporating an "anchor point" in the alignment process
#
# Before we can do any tests, we need to set up the environment, create the factory
# and read in the unaligned sequences.
#
#BEGIN {
# $ENV{CLUSTALDIR} = '/home/peter/clustalw1.8/';
#}
use Getopt::Long;
use Bio::Tools::Run::Alignment::Clustalw;
use Bio::SimpleAlign;
use Bio::AlignIO;
use Bio::SeqIO;
use strict;
# set some default values
my $infile = 't/data/cysprot1a.fa';
my @params = ('quiet' => 1 );
my $do_only = '123'; # string listing examples to be executed. Default is to
# execute all tests (ie 1,2 and 3)
my $param = 'ktuple'; # parameter to be varied in example 1
my $startvalue = 1; # initial value for parameter $param
my $stopvalue = 3; # final value for parameter $param
my $regex = 'W[AT]F'; # regular expression for 'anchoring' alignment in example 3
my $extension = 30; # distance regexp anchor should be extended in each direction
# for local alignment in example 3
my $helpflag = 0; # Flag to show usage info.
# get user options
my @argv = @ARGV; # copy ARGV before GetOptions() massacres it.
&GetOptions("h!" => \$helpflag, "help!" => \$helpflag,
"in=s" => \$infile,
"param=s" => \$param,
"do=s" => \$do_only,
"start=i" => \$startvalue,
"stop=i" => \$stopvalue,
"ext=i" => \$extension,
"regex=s" => \$regex,) ;
if ($helpflag) { &clustalw_usage(); exit 0;}
# create factory & set user-specified global clustalw parameters
foreach my $argv (@argv) {
unless ($argv =~ /^(.*)=>(.*)$/) { next;}
push (@params, $1 => $2);
}
my $factory = Bio::Tools::Run::Alignment::Clustalw->new(@params);
# put unaligned sequences in a Bio::Seq array
my $str = Bio::SeqIO->new(-file=> $infile, '-format' => 'Fasta');
my ($paramvalue, $aln, $subaln, @consensus, $seq_num, $string, $strout, $id);
my @seq_array =();
while ( my $seq = $str->next_seq() ) { push (@seq_array, $seq) ;}
# Do each example that has digit present in variable $do_only
$_= $do_only;
/1/ && &vary_params();
/2/ && &vary_align_order();
/3/ && &anchored_align();
## End of "main"
#################################################
# vary_params(): Example demonstrating varying of clustalw parameter
#
sub vary_params {
print "Beginning parameter-varying example... \n";
# Now we'll create several alignments, 1 for each value of the selected
# parameter. We also compute a simple consensus string for each alignment.
# (In the default case, we vary the "ktuple" parameter, creating 3
# alignments using ktuple values from 1 to 3.)
my $index =0;
for ($paramvalue = $startvalue; $paramvalue < ($stopvalue + 1); $paramvalue++) {
$factory->$param($paramvalue); # set parameter value
print "Performing alignment with $param = $paramvalue \n";
$aln = $factory->align(\@seq_array);
$string = $aln->consensus_string(); # Get consensus of alignment
# convert '?' to 'X' at non-consensus positions
$string =~ s/\?/X/g;
$consensus[$index] = Bio::Seq->new(-id=>"$param=$paramvalue",-seq=>$string);
$index++;
}
# Compare consensus strings for alignments with different $param values by
# making an alignment of the different consensus strings
# $factory->ktuple(1); # set ktuple parameter
print "Performing alignment of $param consensus sequences \n";
$aln = $factory->align(\@consensus);
$strout = Bio::AlignIO->newFh('-format' => 'msf');
print $strout $aln;
return 1;
}
#################################################
# vary_align_order():
#
# For our second example, we'll test the effect of changing the order
# that sequences are added to the alignment
sub vary_align_order {
print "\nBeginning alignment-order-changing example... \n";
@consensus = (); # clear array
for ($seq_num = 0; $seq_num < scalar(@seq_array); $seq_num++) {
my $obj_out = shift @seq_array; # remove one Seq object from array and save
$id = $obj_out->display_id;
# align remaining sequences
print "Performing alignment with sequence $id left out \n";
$subaln = $factory->align(\@seq_array);
# add left-out sequence to subalignment
$aln = $factory->profile_align($subaln,$obj_out);
$string = $aln->consensus_string(); # Get consensus of alignment
# convert '?' to 'X' for non-consensus positions
$string =~ s/\?/X/g;
$consensus[$seq_num] = Bio::Seq->new(-id=>"$id left out",-seq=>$string);
push @seq_array, $obj_out; # return Seq object for next (sub) alignment
}
# Compare consensus strings for alignments created in different orders
# $factory->ktuple(1); # set ktuple parameter
print "\nPerforming alignment of consensus sequences for different reorderings \n";
print "Each consensus is labeled by the sequence which was omitted in the initial alignment\n";
$aln = $factory->align(\@consensus);
$strout = Bio::AlignIO->newFh('-format' => 'msf');
print $strout $aln;
return 1;
}
#################################################
# anchored_align()
#
# For our last example, we'll test a way to perform a local alignment by
# "anchoring" the alignment to a regular expression. This is similar
# to the approach taken in the recent dbclustal program.
# In principle, we could write a script to search for a good regular expression
# to use. Instead, here we'll simply choose one manually after looking at the
# previous alignments.
sub anchored_align {
my @local_array = ();
my @seqs_not_matched = ();
print "\n Beginning anchored-alignment example... \n";
for ($seq_num = 0; $seq_num < scalar(@seq_array); $seq_num++) {
my $seqobj = $seq_array[$seq_num];
my $seq = $seqobj->seq();
my $id = $seqobj->id();
# if $regex is not found in the sequence, save sequence id name and set
# array value =0 for later
unless ($seq =~/$regex/) {
$local_array[$seq_num] = 0;
push (@seqs_not_matched, $id) ;
next;
}
# find positions of start and of subsequence to be aligned
my $match_start_pos = length($`);
my $match_stop_pos = length($`) + length($&);
my $start = ($match_start_pos - $extension) > 1 ?
($match_start_pos - $extension) +1 : 1;
my $stop = ($match_stop_pos + $extension) < length($seq) ?
($match_stop_pos + $extension) : length($seq);
my $string = $seqobj->subseq($start, $stop);
$local_array[$seq_num] = Bio::Seq->new(-id=>$id, -seq=>$string);
}
@local_array = grep $_ , @local_array; # remove array entries with no match
# Perform alignment on the local segments of the sequences which match "anchor"
$aln = $factory->align(\@local_array);
my $consensus = $aln->consensus_string(); # Get consensus of local alignment
if (scalar(@seqs_not_matched) ) {
print " Sequences not matching $regex : @seqs_not_matched \n"
} else {
print " All sequences match $regex : @seqs_not_matched \n"
}
print "Consensus sequence of local alignment: $consensus \n";
return 1;
}
#----------------
sub clustalw_usage {
#----------------
#-----------------------
# Prints usage information for general parameters.
print STDERR <<"QQ_PARAMS_QQ";
Command-line accessible script variables and commands:
-------------------------------
-h : Display this usage info and exit.
-in <str> : File containing input sequences in fasta format (default = $infile) .
-do <str> : String listing examples to be executed. Default is to execute
all tests (ie default = '123')
-param <str> : Parameter to be varied in example 1. Any clustalw parameter
which takes inteer values can be varied (default = 'ktuple')
-start <int> : Initial value for varying parameter in example 1 (default = 1)
-stop <int> : Final value for varying parameter (default = 3)
-regex <str> : Regular expression for 'anchoring' alignment in example 3
(default = $regex)
-ext <int> : Distance regexp anchor should be extended in each direction
for local alignment in example 3 (default = 30)
In addition, any valid Clustalw parameter can be set using the syntax
"parameter=>value" as in "ktuple=>3"
So a typical command lines might be:
> clustalw.pl -param=pairgap -start=2 -stop=3 -do=1 "ktuple=>3"
or
> clustalw.pl -ext=10 -regex='W[AST]F' -do=23 -in='t/cysprot1a.fa'
QQ_PARAMS_QQ
}