Fetching contributors…
Cannot retrieve contributors at this time
891 lines (704 sloc) 26 KB
# BioPerl module for Bio::Tools::Run::Phylo::SLR
# Please direct questions and support issues to <>
# Cared for by Albert Vilella <avilella-at-gmail-dot-com>
# Copyright Albert Vilella
# You may distribute this module under the same terms as perl itself
# POD documentation - main docs before the code
=head1 NAME
Bio::Tools::Run::Phylo::SLR - Wrapper around the SLR program
use Bio::Tools::Run::Phylo::SLR;
use Bio::AlignIO;
use Bio::TreeIO;
use Bio::SimpleAlign;
my $alignio = Bio::AlignIO->new
(-format => 'fasta',
-file => 't/data/219877.cdna.fasta');
my $aln = $alignio->next_aln;
my $treeio = Bio::TreeIO->new
(-format => 'newick', -file => 't/data/219877.tree');
my $tree = $treeio->next_tree;
my $slr = Bio::Tools::Run::Phylo::SLR->new();
# $rc = 1 for success, 0 for errors
my ($rc,$results) = $slr->run();
my $positive_sites = $results->{'positive'};
print "# Site\tNeutral\tOptimal\tOmega\t",
foreach my $positive_site (@$positive_sites) {
$positive_site->[0], "\t",
$positive_site->[1], "\t",
$positive_site->[2], "\t",
$positive_site->[3], "\t",
$positive_site->[4], "\t",
$positive_site->[5], "\t",
$positive_site->[6], "\t",
$positive_site->[7], "\t",
$positive_site->[8], "\t",
This is a wrapper around the SLR program. See for more information.
This module is more about generating the proper ctl file and
will run the program in a separate temporary directory to avoid
creating temp files all over the place.
=head2 Mailing Lists
User feedback is an integral part of the evolution of this and other
Bioperl modules. Send your comments and suggestions preferably to
the Bioperl mailing list. Your participation is much appreciated. - General discussion - About the mailing lists
=head2 Support
Please direct usage questions or support issues to the mailing list:
rather than to the module maintainer directly. Many experienced and
reponsive experts will be able look at the problem and quickly
address it. Please include a thorough description of the problem
with code and data examples if at all possible.
=head2 Reporting Bugs
Report bugs to the Bioperl bug tracking system to help us keep track
of the bugs and their resolution. Bug reports can be submitted via the
=head1 AUTHOR - Albert Vilella
Email avilella-at-gmail-dot-com
Additional contributors names and emails here
The rest of the documentation details each of the object methods.
Internal methods are usually preceded with a _
#' keep my emacs happy
# Let the code begin...
package Bio::Tools::Run::Phylo::SLR;
use strict;
use Bio::Root::Root;
use Bio::AlignIO;
use Bio::TreeIO;
use Bio::SimpleAlign;
use Bio::Tools::Run::WrapperBase;
use Cwd;
use File::Spec;
@ISA = qw(Bio::Root::Root Bio::Tools::Run::WrapperBase);
=head2 Default Values
seqfile [incodon]
File from which to read alignment of codon sequences. The file
should be in PAML format.
treefile [intree]
File from which tree should be read. The tree should be in Nexus
outfile [slr.res]
File to which results are written. If the file already exists, it will
be overwritten.
reoptimise [1]
Should the branch lengths, omega and kappa be reoptimized?
0 - no
1 - yes.
kappa [2.0]
Value for kappa. If 'reoptimise' is specified, the value
given will be used as am initial estimate,
omega [0.1]
Value for omega (dN/dS). If 'reoptimise' is specified, the value
given will be used as an initial estimate.
codonf [0]
How codon frequencies are estimated:
0: F61/F60 Estimates used are the empirical frequencies from the
1: F3x4 The frequencies of nucleotides at each codon position
are estimated from the data and then multiplied together to get the
frequency of observing a given codon. The frequency of stop codons is
set to zero, and all other frequencies scaled appropriately.
2: F1x4 Nucleotide frequencies are estimated from the data
(not taking into account at which position in the codon it occurs).
The nucleotide frequencies are multiplied together to get the frequency
of observing and then corrected for stop codons.
freqtype [0]
How codon frequencies are incorporated into the substitution matrix.
0: q_{ij} = pi_{j} s_{ij}
1: q_{ij} = \sqrt(pi_j/pi_i) s_{ij}
2: q_{ij} = \pi_{n} s_{ij}, where n is the nucleotide that the
subsitution is to.
3: q_{ij} = s_{ij} / pi_i
Option 0 is the tradition method of incorporating equilibrium frequencies
into subsitution matrices (Felsenstein 1981; Goldman and Yang, 1994)
Option 1 is described by Goldman and Whelan (2002), in this case with the
additional parameter set to 0.5.
Option 2 was suggested by Muse and Gaut (1994).
Option 3 is included as an experiment, originally suggested by Bret Larget.
it does not appear to describe evolution very successfully and should not
be used for analyses.
Kosakovsky-Pond has repeatedly stated that he finds incorporating codon
frequencies in the manner of option 2 to be superior to option 0. We find
that option 1 tends to perform better than either of these options.
positive_only [0]
If only positively selected sites are of interest, set this to "1".
Calculation will be slightly faster, but information about sites under
purifying selection is lost.
gencode [universal]
Which genetic code to use when determining whether a given mutation
is synonymous or nonsynonymous. Currently only "universal" and
"mammalian" mitochondrial are supported.
nucleof [0]
Allow for empirical exchangabilities for nucleotide substitution.
0: No adjustment. All nucleotides treated the same, modulo
transition / transversion.
1: The rate at which a substitution caused a mutation from nucleotide
a to nucleotide b is adjust by a constant N_{ab}. This adjustment is
in addition to other adjustments (e.g. transition / transversion or
base frequencies).
aminof [0]
Incorporate amino acid similarity parameters into substitution matrix,
adjusting omega for a change between amino acid i and amino acid j.
A_{ij} is a symmetric matrix of constants representing amino acid
0: Constant omega for all amino acid changes
1: omega_{ij} = omega^{A_{ij}}
2: omega_{ij} = a_{ij} log(omega) / [ 1 - exp(-a_{ij} log(omega)) ]
Option 1 has the same form as the original codon subsitution model
proposed by Goldman and Yang (but with potentially different
Option 2 has a more population genetic derivtion, with omega being
interpreted as the ratio of fixation probabilities.
nucfile [nuc.dat]
If nucleof is non-zero, read nucleotide substitution constants from
nucfile. If this file does not exist, hard coded constants are used.
aminofile [amino.dat]
If aminof is non-zero, read amino acid similarity constants from
aminofile. If this file does not exist, hard coded constants are used.
timemem [0]
Print summary of real time and CPU time used. Will eventually print
summary of memory use as well.
ldiff [3.841459]
Twice log-likelihood difference used as a threshold for calculating
support (confidence) intervals for sitewise omega estimates. This
value should be the quantile from a chi-square distribution with one
degree of freedom corresponding to the support required.
E.g. qchisq(0.95,1) = 3.841459
0.4549364 = 50% support
1.323304 = 75% support
2.705543 = 90% support
3.841459 = 95% support
6.634897 = 99% support
7.879439 = 99.5% support
10.82757 = 99.9% support
paramin []
If not blank, read in parameters from file given by the argument.
paramout []
If not blank, write out parameter estimates to file given.
skipsitewise [0]
Skip sitewise estimation of omega. Depending on other options given,
either calculate maximum likelihood or likelihood fixed at parameter
values given.
seed [0]
Seed for random number generator. If seed is 0, then previously
produced seed file (~/.rng64) is used. If this does not exist, the
random number generator is initialised using the clock.
saveseed [1]
If non-zero, save finial seed in file (~/.rng64) to be used as initial
seed in future runs of program.
=head2 Results Format
Results file (default: slr.res)
Results are presented in nine columns
Number of sites in alignment
(minus) Log-probability of observing site given that it was
evolving neutrally (omega=1)
(minus) Log-probability of observing site given that it was
evolving at the optimal value of omega.
The value of omega which maximizes the log-probability of observing
Log-likelihood ratio statistic for non-neutral selection (or
positive selection if the positive_only option is set to 1).
LRT_Stat = 2 * (Neutral-Optimal)
P-value for non-neutral (or positive) selection at a site,
unadjusted for multiple comparisons.
Adj. Pval
P-value for non-neutral (or positive) selection at a site, after
adjusting for multiple comparisons using the Hochberg procedure
(see the file "MultipleComparisons.txt" in the doc directory).
A simple visual guide to the result. Sites detected as having been
under positive selection are marked with a '+', sites under
purifying selection are marked with '-'. The number of symbols
Number symbols Threshold
1 95%
2 99%
3 95% after adjustment
4 99% after adjustment
Occasionally the result may also contain an exclamation mark. This
indicates that the observation at a site is not significantly
different from random (equivalent to infinitely strong positive
selection). This may indicate that the alignment at that site is bad
The following events are flagged:
Synonymous All codons at a site code for the same amino
Single character Only one sequence at the site is ungapped,
the result of a recent insertion for example.
All gaps All sequences at a site contain a gap
Sites marked "Single character" or "All gaps" are not counted
towards the number of sites for the purposes of correcting for
multiple comparisons since it is not possible to detect selection
from none or one observation under the assumptions made by the
sitewise likelihood ratio test.
#' keep my emacs happy
$PROGRAMNAME = 'Slr_Linux_static';
if ($^O =~ /darwin/i) {
$PROGRAMNAME = 'Slr_osx';
} elsif ($^O =~ /mswin/i) {
$PROGRAMNAME = 'Slr_windows.exe';
if( defined $ENV{'SLRDIR'} ) {
$PROGRAM = Bio::Root::IO->catfile($ENV{'SLRDIR'},$PROGRAMNAME). ($^O =~ /mswin/i ?'_windows.exe':'');;
# valid values for parameters, the default one is always
# the first one in the array
# example file provided with the package
'outfile' => 'slr.res',
'reoptimise' => [ 1,0],
'kappa' => '2.0',
'omega' => '0.1',
'codonf' => [ 0, 1,2],
'freqtype' => [ 0, 1,2,3],
'positive_only' => [ 0, 1],
'gencode' => [ "universal", "mammalian"],
'nucleof' => [ 0, 1],
'aminof' => [ 0, 1,2],
'nucfile' => '',
'aminofile' => '',
'timemem' => [ 0, 1],
'ldiff' => [ 3.841459, 0.4549364,1.323304,2.705543,6.634897,7.879439,10.82757],
'paramin' => '',
'paramout' => '',
'skipsitewise' => [ 0, 1],
'seed' => [0],
'saveseed' => [ 1, 0]
=head2 program_name
Title : program_name
Usage : $factory->program_name()
Function: holds the program name
Returns: string
Args : None
sub program_name {
=head2 program_dir
Title : program_dir
Usage : ->program_dir()
Function: returns the program directory, obtained from ENV variable.
Returns: string
Args :
sub program_dir {
return Bio::Root::IO->catfile($ENV{SLRDIR}) if $ENV{SLRDIR};
=head2 new
Title : new
Usage : my $obj = Bio::Tools::Run::Phylo::SLR->new();
Function: Builds a new Bio::Tools::Run::Phylo::SLR object
Returns : Bio::Tools::Run::Phylo::SLR
Args : -alignment => the Bio::Align::AlignI object
-save_tempfiles => boolean to save the generated tempfiles and
NOT cleanup after onesself (default FALSE)
-tree => the Bio::Tree::TreeI object
-params => a hashref of SLR parameters (all passed to set_parameter)
-executable => where the SLR executable resides
See also: L<Bio::Tree::TreeI>, L<Bio::Align::AlignI>
sub new {
my($class,@args) = @_;
my $self = $class->SUPER::new(@args);
my ($aln, $tree, $st, $params, $exe,
$ubl) = $self->_rearrange([qw(ALIGNMENT TREE SAVE_TEMPFILES
defined $aln && $self->alignment($aln);
defined $tree && $self->tree($tree);
defined $st && $self->save_tempfiles($st);
defined $exe && $self->executable($exe);
if( defined $params ) {
if( ref($params) !~ /HASH/i ) {
$self->warn("Must provide a valid hash ref for parameter -FLAGS");
} else {
map { $self->set_parameter($_, $$params{$_}) } keys %$params;
return $self;
=head2 prepare
Title : prepare
Usage : my $rundir = $slr->prepare($aln);
Function: prepare the SLR analysis using the default or updated parameters
the alignment parameter must have been set
Returns : value of rundir
Args : L<Bio::Align::AlignI> object,
L<Bio::Tree::TreeI> object
sub prepare{
my ($self,$aln,$tree) = @_;
unless ( $self->save_tempfiles ) {
# brush so we don't get plaque buildup ;)
$tree = $self->tree unless $tree;
$aln = $self->alignment unless $aln;
if( ! $aln ) {
$self->warn("must have supplied a valid alignment file in order to run SLR");
return 0;
if( ! $tree ) {
$self->warn("must have supplied a valid tree file in order to run SLR");
return 0;
my ($tempdir) = $self->tempdir();
my ($tempseqFH,$tempseqfile);
# Reorder the alignment according to the tree
my $ct = 1;
my %order;
foreach my $node ($tree->get_leaf_nodes) {
$order{$node->id_output} = $ct++;
my @seq; my @ids;
foreach my $seq ( $aln->each_seq() ) {
push @seq, $seq;
push @ids, $seq->display_id;
# use the map-sort-map idiom:
my @sorted = map { $_->[1] } sort { $a->[0] <=> $b->[0] } map { [$order{$_->id()}, $_] } @seq;
my $sorted_aln = Bio::SimpleAlign->new();
foreach (@sorted) {
# Rename the leaf nodes in the tree from 1 to n
$ct = 1;
foreach my $node ($tree->get_leaf_nodes) {
($tempseqFH,$tempseqfile) = $self->io->tempfile
('-dir' => $tempdir,
UNLINK => ($self->save_tempfiles ? 0 : 1));
my $alnout = Bio::AlignIO->new('-format' => 'phylip',
'-fh' => $tempseqFH,
'-interleaved' => 0,
'-idlinebreak' => 1,
'-idlength' => $MINNAMELEN > $aln->maxdisplayname_length() ? $MINNAMELEN : $aln->maxdisplayname_length() +1);
undef $alnout;
my ($temptreeFH,$temptreefile);
($temptreeFH,$temptreefile) = $self->io->tempfile
('-dir' => $tempdir,
UNLINK => ($self->save_tempfiles ? 0 : 1));
my $treeout = Bio::TreeIO->new('-format' => 'newick',
'-fh' => $temptreeFH);
# We need to add a line with the num of leaves ($ct-1) and the
# num of trees (1)
$treeout->_print(sprintf("%d 1\n",($ct-1)));
# now let's print the ctl file.
# many of the these programs are finicky about what the filename is
# and won't even run without the properly named file.
my ($treevolume,$treedirectories,$treefile) = File::Spec->splitpath( $temptreefile );
my ($alnvolume,$alndirectories,$alnfile) = File::Spec->splitpath( $tempseqfile );
my $slr_ctl = "$tempdir/slr.ctl";
open(SLR, ">$slr_ctl") or $self->throw("cannot open $slr_ctl for writing");
print SLR "seqfile\: $alnfile\n";
print SLR "treefile\: $treefile\n";
my $outfile = $self->outfile_name;
print SLR "outfile\: $outfile\n";
my %params = $self->get_parameters;
while( my ($param,$val) = each %params ) {
next if $param eq 'outfile';
print SLR "$param\: $val\n";
return $tempdir;
=head2 run
Title : run
Usage : my ($rc,$parser) = $slr->run($aln,$tree);
Function: run the SLR analysis using the default or updated parameters
the alignment parameter must have been set
Returns : Return code, L<Bio::Tools::Phylo::SLR>
Args : L<Bio::Align::AlignI> object,
L<Bio::Tree::TreeI> object
sub run {
my ($self) = shift;;
my $outfile = $self->outfile_name;
my $tmpdir = $self->prepare(@_);
#my ($rc,$parser) = (1);
my ($rc,$results) = (1);
my $cwd = cwd();
my $exit_status;
my $slrexe = $self->executable();
$self->throw("unable to find or run executable for SLR") unless $slrexe && -e $slrexe && -x _;
my $run;
open($run, "$slrexe |") or $self->throw("Cannot open exe $slrexe");
my @output = <$run>;
$exit_status = close($run);
if( (grep { /\berr(or)?: /io } @output) || !$exit_status) {
$self->warn("There was an error - see error_string for the program output");
$rc = 0;
eval {
open RESULTS, "$tmpdir/$outfile" or die "couldnt open results file: $!\n";
my $okay = 0;
my $sites;
my $type = 'default';
while (<RESULTS>) {
chomp $_;
if ( /^\#/ ) {next;}
if ( /\!/ ) {$type = 'random';} # random is last
elsif ( /\+/ ) {$type = 'positive';}
elsif ( /\-\s+/ ) {$type = 'negative';}
elsif ( /Constant/ ) {$type = 'constant';}
elsif ( /All gaps/ ) {$type = 'all_gaps';}
elsif ( /Single character/ ) {$type = 'single_character';}
elsif ( /Synonymous/ ) {$type = 'synonymous';}
else {$type = 'default'}
if ( /^\s+(\d+)\s+(\S+)\s+(\S+)\s+(\S+)\s+(\S+)\s+(\S+)\s+(\S+)\s+(\S+)\s+(\S+)/ ) {
push @{$sites->{$type}}, [$1,$2,$3,$4,$5,$6,$7,$8,$9];
} else {
$results = $sites;
close RESULTS;
# TODO: we could have a proper parser object
# $parser = Bio::Tools::Phylo::SLR->new(-file => "$tmpdir/$outfile",
# -dir => "$tmpdir");
if( $@ ) {
# return ($rc,$parser);
return ($rc,$results);
=head2 error_string
Title : error_string
Usage : $obj->error_string($newval)
Function: Where the output from the last analysus run is stored.
Returns : value of error_string
Args : newvalue (optional)
sub error_string{
my ($self,$value) = @_;
if( defined $value) {
$self->{'error_string'} = $value;
return $self->{'error_string'};
=head2 alignment
Title : alignment
Usage : $slr->align($aln);
Function: Get/Set the L<Bio::Align::AlignI> object
Returns : L<Bio::Align::AlignI> object
Args : [optional] L<Bio::Align::AlignI>
Comment : We could potentially add support for running directly on a file
but we shall keep it simple
See also: L<Bio::SimpleAlign>
sub alignment{
my ($self,$aln) = @_;
if( defined $aln ) {
if( -e $aln ) {
$self->{'_alignment'} = $aln;
} elsif( !ref($aln) || ! $aln->isa('Bio::Align::AlignI') ) {
$self->warn("Must specify a valid Bio::Align::AlignI object to the alignment function not $aln");
return undef;
} else {
$self->{'_alignment'} = $aln;
return $self->{'_alignment'};
=head2 tree
Title : tree
Usage : $slr->tree($tree, %params);
Function: Get/Set the L<Bio::Tree::TreeI> object
Returns : L<Bio::Tree::TreeI>
Args : [optional] $tree => L<Bio::Tree::TreeI>,
Comment : We could potentially add support for running directly on a file
but we shall keep it simple
See also: L<Bio::Tree::Tree>
sub tree {
my ($self, $tree, %params) = @_;
if( defined $tree ) {
if( ! ref($tree) || ! $tree->isa('Bio::Tree::TreeI') ) {
$self->warn("Must specify a valid Bio::Tree::TreeI object to the alignment function");
$self->{'_tree'} = $tree;
return $self->{'_tree'};
=head2 get_parameters
Title : get_parameters
Usage : my %params = $self->get_parameters();
Function: returns the list of parameters as a hash
Returns : associative array keyed on parameter names
Args : none
sub get_parameters{
my ($self) = @_;
# we're returning a copy of this
return %{ $self->{'_slrparams'} };
=head2 set_parameter
Title : set_parameter
Usage : $slr->set_parameter($param,$val);
Function: Sets a SLR parameter, will be validated against
the valid values as set in the %VALIDVALUES class variable.
The checks can be ignored if one turns off param checks like this:
Returns : boolean if set was success, if verbose is set to -1
then no warning will be reported
Args : $param => name of the parameter
$value => value to set the parameter to
See also: L<no_param_checks()>
sub set_parameter{
my ($self,$param,$value) = @_;
unless (defined $self->{'no_param_checks'} && $self->{'no_param_checks'} == 1) {
if ( ! defined $VALIDVALUES{$param} ) {
$self->warn("unknown parameter $param will not be set unless you force by setting no_param_checks to true");
return 0;
if ( ref( $VALIDVALUES{$param}) =~ /ARRAY/i &&
scalar @{$VALIDVALUES{$param}} > 0 ) {
unless ( grep { $value eq $_ } @{ $VALIDVALUES{$param} } ) {
$self->warn("parameter $param specified value $value is not recognized, please see the documentation and the code for this module or set the no_param_checks to a true value");
return 0;
$self->{'_slrparams'}->{$param} = $value;
return 1;
=head2 set_default_parameters
Title : set_default_parameters
Usage : $slr->set_default_parameters(0);
Function: (Re)set the default parameters from the defaults
(the first value in each array in the
%VALIDVALUES class variable)
Returns : none
Args : boolean: keep existing parameter values
sub set_default_parameters{
my ($self,$keepold) = @_;
$keepold = 0 unless defined $keepold;
while( my ($param,$val) = each %VALIDVALUES ) {
# skip if we want to keep old values and it is already set
next if( defined $self->{'_slrparams'}->{$param} && $keepold);
if(ref($val)=~/ARRAY/i ) {
$self->{'_slrparams'}->{$param} = $val->[0];
} else {
$self->{'_slrparams'}->{$param} = $val;
=head1 Bio::Tools::Run::WrapperBase methods
=head2 no_param_checks
Title : no_param_checks
Usage : $obj->no_param_checks($newval)
Function: Boolean flag as to whether or not we should
trust the sanity checks for parameter values
Returns : value of no_param_checks
Args : newvalue (optional)
sub no_param_checks{
my ($self,$value) = @_;
if( defined $value) {
$self->{'no_param_checks'} = $value;
return $self->{'no_param_checks'};
=head2 save_tempfiles
Title : save_tempfiles
Usage : $obj->save_tempfiles($newval)
Returns : value of save_tempfiles
Args : newvalue (optional)
=head2 outfile_name
Title : outfile_name
Usage : my $outfile = $slr->outfile_name();
Function: Get/Set the name of the output file for this run
(if you wanted to do something special)
Returns : string
Args : [optional] string to set value to
sub outfile_name {
my $self = shift;
if( @_ ) {
return $self->{'_slrparams'}->{'outfile'} = shift @_;
unless (defined $self->{'_slrparams'}->{'outfile'}) {
$self->{'_slrparams'}->{'outfile'} = 'out.res';
return $self->{'_slrparams'}->{'outfile'};
=head2 tempdir
Title : tempdir
Usage : my $tmpdir = $self->tempdir();
Function: Retrieve a temporary directory name (which is created)
Returns : string which is the name of the temporary directory
Args : none
=head2 cleanup
Title : cleanup
Usage : $slr->cleanup();
Function: Will cleanup the tempdir directory after an SLR run
Returns : none
Args : none
=head2 io
Title : io
Usage : $obj->io($newval)
Function: Gets a L<Bio::Root::IO> object
Returns : L<Bio::Root::IO>
Args : none
my $self= shift;
unless ( $self->save_tempfiles ) {