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extractFromGenomeByGFF.pl
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extractFromGenomeByGFF.pl
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#!/usr/bin/perl
#
# INGLÊS/ENGLISH
# This program is distributed in the hope that it will be useful,
# but WITHOUT ANY WARRANTY; without even the implied warranty of
# MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the
# GNU General Public License for more details.
# http://www.gnu.org/copyleft/gpl.html
#
#
# PORTUGUÊS/PORTUGUESE
# Este programa é distribuído na expectativa de ser útil aos seus
# usuários, porém NÃO TEM NENHUMA GARANTIA, EXPLÍCITAS OU IMPLÍCITAS,
# COMERCIAIS OU DE ATENDIMENTO A UMA DETERMINADA FINALIDADE. Consulte
# a Licença Pública Geral GNU para maiores detalhes.
# http://www.gnu.org/copyleft/gpl.html
#
# Copyright (C) 2012 Universidade de São Paulo
#
# Universidade de São Paulo
# Laboratório de Biologia do Desenvolvimento de Abelhas
# Núcleo de Bioinformática (LBDA-BioInfo)
#
# Daniel Guariz Pinheiro
# http://zulu.fmrp.usp.br/bioinfo
#
# $Id$
=head1 NAME
extractFromGenomeByGFF.pl
=head1 SYNOPSIS
perl extractFromGenomeByGFF.pl -g /data/files/Amel_4.5/BeeBase/ -f amel_OGSv3.1_with_UTR.gff3 -o ./output \
-p OGS_
=head1 ABSTRACT
Extract sequences of 5' UTR, 3' UTR and CDS of mRNA sequences from chromosome files in FASTA format using
GFF coordinates. The global statistics are displayed in STDERR output.
=head1 DESCRIPTION
Arguments:
-h/--help Help
-l/--level Log level [Default: FATAL]
OFF
FATAL
ERROR
WARN
INFO
DEBUG
TRACE
ALL
-g/--genome Genome directory
-f/--gff GFF file
-o/--outdir Output directory
-p/--outprefix Output filenames' prefix
=head1 AUTHOR
Daniel Guariz Pinheiro E<lt>[email protected]<gt>
Copyright (c) 2012 Universidade de São Paulo
=head1 LICENSE
GNU General Public License
http://www.gnu.org/copyleft/gpl.html
=cut
use strict;
use warnings;
use Readonly;
use Getopt::Long;
#use Bio::Factory::SeqAnalysisParserFactory;
use Bio::Tools::GFF;
use Bio::SeqIO;
use Bio::Seq;
use vars qw/$LOGGER/;
INIT {
use Log::Log4perl qw/:easy/;
Log::Log4perl->easy_init($FATAL);
$LOGGER = Log::Log4perl->get_logger($0);
}
my ($level, $gfffile, $genomedir, $outdir, $outprefix, $genomefile, $statsfile, $threepminsize);
Usage("Too few arguments") if $#ARGV < 0;
GetOptions( "h|?|help" => sub { &Usage(); },
"l|level=s"=> \$level,
"d|genomedir=s"=>\$genomedir,
"g|genomefile=s"=>\$genomefile,
"s|statsfile=s"=>\$statsfile,
"f|gff=s"=>\$gfffile,
"o|outdir=s"=>\$outdir,
"p|outprefix=s"=>\$outprefix,
"m|threepminsize=i"=>\$threepminsize
) or &Usage();
$threepminsize||=0;
if ($level) {
my %LEVEL = (
'OFF' =>$OFF,
'FATAL' =>$FATAL,
'ERROR' =>$ERROR,
'WARN' =>$WARN,
'INFO' =>$INFO,
'DEBUG' =>$DEBUG,
'TRACE' =>$TRACE,
'ALL' =>$ALL);
$LOGGER->logdie("Wrong log level ($level). Choose one of: ".join(', ', keys %LEVEL)) unless (exists $LEVEL{$level});
Log::Log4perl->easy_init($LEVEL{$level});
}
use File::Temp qw/ tempfile tempdir /;
$LOGGER->logdie('Missing genome directory or genome file') if ((!$genomedir)&&(!$genomefile));
if ($genomefile) {
$LOGGER->logdie('Wrong genome file ('.$genomefile.')') unless (-e $genomefile);
my $in = Bio::SeqIO->new(
-file => $genomefile,
-format => 'fasta'
);
if ($genomedir) {
unless (-e $genomedir) {
mkdir($genomedir);
$LOGGER->logdie('Cannot create genome directory ('.$genomedir.')') unless (-d $genomedir);
}
}
else {
$genomedir = tempdir( CLEANUP => 1 );
}
my @f = glob("$genomedir/*.fa");
if ( scalar(@f) < 1) {
while ( my $seq = $in->next_seq() ) {
my $out = Bio::SeqIO->new(
-file => '>'.$genomedir.'/'.$seq->display_id().'.fa',
-format => 'fasta'
);
$out->write_seq($seq);
}
}
}
else {
if ($genomedir) {
$LOGGER->logdie('Wrong genome directory ('.$genomedir.')') unless (-d $genomedir);
}
}
$LOGGER->logdie('Missing gff file') unless ($gfffile);
$LOGGER->logdie('Wrong gff file ('.$gfffile.')') unless (-e $gfffile);
if ($outdir) {
$LOGGER->logdie('Wrong output directory ('.$outdir.')') unless (-d $outdir);
}
else {
$outdir='.';
}
$outprefix||='out_';
my %feature;
my %parent;
my $gffio = Bio::Tools::GFF->new(-file =>$gfffile, -gff_version => 3);
my $feat;
# loop over the input stream
my $c = 0;
my %chromosome;
my $idcounter = 0;
my %allgene;
while ($feat = $gffio->next_feature()) {
$c++;
die "Not found ".$feat->seq_id()." on $genomedir" unless (-e "$genomedir/".$feat->seq_id().'.fa');
my ($id,$parent);
if ($feat->has_tag('ID')) {
($id) = $feat->get_tag_values('ID');
}
else {
($id) = 'UNK'.$idcounter++;
}
unless (exists $feature{$id}) {
$feature{$id} = { 'primary_tag' => $feat->primary_tag(),
'child' => undef };
push(@{$chromosome{ $feat->seq_id() }}, $id);
}
if ($feat->has_tag('Parent')) {
($parent) = $feat->get_tag_values('Parent');
if ($id) {
$parent{$id} = $parent unless (exists $parent{$id});
}
push(@{$feature{$parent}->{'child'}->{ $feat->primary_tag() }}, $feat);
}
else {
$allgene{ $feat->seq_id() }->{$id} = $feat;
}
}
my %out;
my %count = (
'scaffold' => 0,
'gene' => 0,
'mRNA' => 0,
'5pCDS3pmRNA' => 0,
'5pCDSmRNA' => 0,
'CDS3pmRNA' => 0,
'CDSmRNA' => 0,
'ncRNA' => 0,
'byGroup' => {}
);
foreach my $chr ( keys %allgene ) {
my ($group) = $chr=~/^(Group(?:\d+|Un\d*)|MT)/i;
@{ $count{'byGroup'}->{$group} }{ 'scaffold', 'gene', 'mRNA', '5pCDS3pmRNA', '5pCDSmRNA', 'CDS3pmRNA', 'CDSmRNA', 'ncRNA' } = (0, 0, 0, 0, 0, 0, 0, 0) unless (exists $count{'byGroup'}->{$group});
$count{'scaffold'}++;
$count{'byGroup'}->{$group}->{'scaffold'}++;
my $in = Bio::SeqIO->new(
-file => "$genomedir/" . $chr . '.fa',
-format => 'fasta'
);
my $seq = $in->next_seq();
foreach my $id ( keys %{$allgene{ $chr } } ) {
# gene
if (!exists $parent{$id}) {
# print $id,"\t",$feature{$id}->{'primary_tag'},"\n";
$count{'gene'}++;
$count{'byGroup'}->{$group}->{'gene'}++;
# print STDERR "GENE: $id","\n";
foreach my $mRNAfeat (@{ $feature{$id}->{'child'}->{ 'mRNA' } } ) {
my ($mRNAid) = $mRNAfeat->get_tag_values('ID');
# print STDERR "\t",$mRNAid,"\n";
my $strseq='';
{
my @fe;
if ($mRNAfeat->strand() == -1) {
@fe = sort { $b->start() <=> $a->start() } @{ $feature{$mRNAid}->{'child'}->{'exon'} };
}
elsif ($mRNAfeat->strand() == 1) {
@fe = sort { $a->start() <=> $b->start() } @{ $feature{$mRNAid}->{'child'}->{'exon'} };
}
else {
$LOGGER->logdie("Cannot found strand for $mRNAid");
}
foreach my $f ( @fe ) {
#print "\t",$f->primary_tag(),"\n";
#print "\t",$seq->subseq($f->location()),"\n";
$strseq.=$seq->subseq($f->location());
}
}
my $extseq='';
if ($threepminsize>0) {
my ($ts, $te) = ($mRNAfeat->start(), $mRNAfeat->end());
if ($mRNAfeat->strand() == -1) {
if ($ts > $threepminsize) {
$extseq = $seq->trunc($ts-1-($threepminsize-1), $ts-1)->revcom()->seq();
}
else {
if ($ts>1) {
$extseq = $seq->trunc(1, $ts-1)->revcom()->seq();
}
}
}
elsif ($mRNAfeat->strand() == 1) {
if ( ($te+1+$threepminsize) <= $seq->length() ) {
$extseq = $seq->trunc($te+1, $te+1+($threepminsize-1))->seq();
}
else {
if ( $te < $seq->length() ) {
$extseq = $seq->trunc($te+1, $seq->length())->seq();
}
}
}
else {
$LOGGER->logdie("Cannot found strand for $mRNAid");
}
}
my $smallest3plen = $threepminsize;
if (exists $feature{$mRNAid}->{'child'}->{'CDS'}) {
my %eachCDS;
{
my @allcds = @{ $feature{$mRNAid}->{'child'}->{'CDS'} };
foreach my $cf (@allcds) {
my ($CDSid);
if ($cf->has_tag('ID')) {
($CDSid) = $cf->get_tag_values('ID');
$CDSid=~s/-\d+$//;
}
else {
# OGS (each mRNA has only one CDS)
($CDSid) = $cf->get_tag_values('Parent');
$CDSid=~s/-R([ABC])/-P$1/;
}
my ($CDSgroup) = $CDSid=~/^([^#]+)/;
push(@{$eachCDS{ $CDSgroup }}, $cf);
}
}
foreach my $CDSgroup (keys %eachCDS) {
my @fc;
if ($mRNAfeat->strand() == -1) {
@fc = sort { $b->start() <=> $a->start() } @{ $eachCDS{ $CDSgroup } };
}
elsif ($mRNAfeat->strand() == 1) {
@fc = sort { $a->start() <=> $b->start() } @{ $eachCDS{ $CDSgroup } };
}
else {
$LOGGER->logdie("Cannot found strand for $mRNAid");
}
my $cdsseq='';
foreach my $f ( @fc ) {
$cdsseq.=$seq->subseq($f->location());
}
{
my $seqobj = Bio::Seq->new(
-seq => $cdsseq,
-display_id => 'CDSmRNA'.'|'.$CDSgroup.'|'.$mRNAid.'|'.$chr,
-description => ''
);
unless (exists $out{'CDSmRNA'}) {
$out{'CDSmRNA'} = Bio::SeqIO->new(
-file => ">$outdir/$outprefix".'CDSmRNA'.'.fa',
-format => 'fasta'
);
}
$out{'CDSmRNA'}->write_seq($seqobj);
}
if ($strseq=~/(.*)$cdsseq(.*)/) {
my ($fivep,$threep) = ($1,$2);
if ($fivep) {
my $seqobj = Bio::Seq->new(
-seq => $fivep,
-display_id => 'UTR5p'.'|'.$CDSgroup.'|'.$mRNAid.'|'.$chr,
-description => ''
);
unless (exists $out{'UTR5p'}) {
$out{'UTR5p'} = Bio::SeqIO->new(
-file => ">$outdir/$outprefix".'UTR5p'.'.fa',
-format => 'fasta'
);
}
$out{'UTR5p'}->write_seq($seqobj);
}
$smallest3plen = length($threep);
my $ext = 0;
if (length($extseq)>0) {
my $lenthreep = (($threep) ? length($threep) : 0);
if (length($extseq)>$lenthreep) {
$ext = length($extseq)-$lenthreep;
}
$threep.=substr($extseq,0, $ext);
}
if ($threep) {
my $seqobj = Bio::Seq->new(
-seq => $threep,
-display_id => 'UTR3p'.(($ext) ? '-'.$ext : '').'|'.$CDSgroup.'|'.$mRNAid.'|'.$chr,
-description => ''
);
unless (exists $out{'UTR3p'}) {
$out{'UTR3p'} = Bio::SeqIO->new(
-file => ">$outdir/$outprefix".'UTR3p'.'.fa',
-format => 'fasta'
);
}
$out{'UTR3p'}->write_seq($seqobj);
}
if ($fivep) {
if ($threep) {
$count{'5pCDS3pmRNA'}++;
$count{'byGroup'}->{$group}->{'5pCDS3pmRNA'}++;
}
else {
$count{'5pCDSmRNA'}++;
$count{'byGroup'}->{$group}->{'5pCDSmRNA'}++;
}
}
elsif ($threep) {
if ($fivep) {
$count{'5pCDS3pmRNA'}++;
$count{'byGroup'}->{$group}->{'5pCDS3pmRNA'}++;
}
else {
$count{'CDS3pmRNA'}++;
$count{'byGroup'}->{$group}->{'CDS3pmRNA'}++;
}
}
else {
$count{'CDSmRNA'}++;
$count{'byGroup'}->{$group}->{'CDSmRNA'}++;
}
#print $CDSgroup."\t".$fivep."\n";
#print $CDSgroup."\t".$threep."\n";
}
else {
$LOGGER->logdie("Something wrong! Not match CDS ($CDSgroup) into mRNA ($mRNAid)");
}
}
{
my $ext = 0;
my $threepcomp = '';
if (length($extseq)>0) {
my $lenthreep = $smallest3plen;
if (length($extseq)>$lenthreep) {
$ext = length($extseq)-$lenthreep;
}
$threepcomp.=substr($extseq,0, $ext);
}
$count{'mRNA'}++;
$count{'byGroup'}->{$group}->{'mRNA'}++;
my $seqobj = Bio::Seq->new(
-seq => $strseq.$threepcomp,
-display_id => 'mRNA'.(($ext) ? '-'.$ext : '').'|'.$mRNAid.'|'.$parent{$mRNAid}.'|'.$chr,
-description => ''
);
unless (exists $out{'mRNA'}) {
$out{'mRNA'} = Bio::SeqIO->new(
-file => ">$outdir/$outprefix".'mRNA'.'.fa',
-format => 'fasta'
);
}
$out{'mRNA'}->write_seq($seqobj);
}
}
else {
$count{'ncRNA'}++;
$count{'byGroup'}->{$group}->{'ncRNA'}++;
my $seqobj = Bio::Seq->new(
-seq => $strseq,
-display_id => 'ncRNA'.'|'.$mRNAid.'|'.$parent{$mRNAid}.'|'.$chr,
-description => ''
);
unless (exists $out{'ncRNA'}) {
$out{'ncRNA'} = Bio::SeqIO->new(
-file => ">$outdir/$outprefix".'ncRNA'.'.fa',
-format => 'fasta'
);
}
$out{'ncRNA'}->write_seq($seqobj);
}
}
}
}
}
if ($statsfile) {
open(STATS, ">", $statsfile) or $LOGGER->logdie("$!");
print STATS "
*** GENERAL REPORT ***
Nº of scaffolds scanned.: $count{'scaffold'}
Nº of genes found.......: $count{'gene'}
Nº of mRNAs found.......: $count{'mRNA'}
5p CDS 3p...........: $count{'5pCDS3pmRNA'}
5p CDS..............: $count{'5pCDSmRNA'}
CDS 3p..............: $count{'CDS3pmRNA'}
CDS.................: $count{'CDSmRNA'}
Nº of ncRNAs found......: $count{'ncRNA'}
";
print STATS "
*** REPORT BY GROUP ***
";
foreach my $group (sort {$a cmp $b} keys %{$count{'byGroup'}} ) {
print STATS "
* $group
Nº of scaffolds scanned.: $count{'byGroup'}->{$group}->{'scaffold'}
Nº of genes found.......: $count{'byGroup'}->{$group}->{'gene'}
Nº of mRNAs found.......: $count{'byGroup'}->{$group}->{'mRNA'}
5p CDS 3p...........: $count{'byGroup'}->{$group}->{'5pCDS3pmRNA'}
5p CDS..............: $count{'byGroup'}->{$group}->{'5pCDSmRNA'}
CDS 3p..............: $count{'byGroup'}->{$group}->{'CDS3pmRNA'}
CDS.................: $count{'byGroup'}->{$group}->{'CDSmRNA'}
Nº of ncRNAs found......: $count{'byGroup'}->{$group}->{'ncRNA'}
";
}
close(STATS);
}
# Subroutines
sub Usage {
my ($msg) = @_;
Readonly my $USAGE => <<"END_USAGE";
Daniel Guariz Pinheiro (dgpinheiro\@gmail.com)
(c)2012 Universidade de São Paulo
Usage
$0 [-h/--help] [-l/--level <LEVEL>]
Argument(s)
-h --help Help
-l --level Log level [Default: FATAL]
-g --genomefile Genome file
-d --genomedir Genome directory
-f --gff GFF file
-o --outdir Output directory
-p --outprefix Output filenames' prefix
-s --statsfile Output Statistics file
-m --threepminsize 3' minimum size (extends if possible) [Default: 0]
END_USAGE
print STDERR "\nERR: $msg\n\n" if $msg;
print STDERR qq[$0 ] . q[$Revision$] . qq[\n];
print STDERR $USAGE;
exit(1);
}