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LTRPipeline
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LTRPipeline
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#!/usr/bin/perl
##---------------------------------------------------------------------------##
## File:
## @(#) LTRPipeline
## Author:
## Robert M. Hubley rhubley@systemsbiology.org
## Description:
## This is a port of Jullien Michelle Flynn's ensemble pipeline
## involving LTRDetector, LTR_retreiver, MAFFT, NINJA and CD-HIT
##
#******************************************************************************
#* This software is provided ``AS IS'' and any express or implied *
#* warranties, including, but not limited to, the implied warranties of *
#* merchantability and fitness for a particular purpose, are disclaimed. *
#* In no event shall the authors or the Institute for Systems Biology *
#* liable for any direct, indirect, incidental, special, exemplary, or *
#* consequential damages (including, but not limited to, procurement of *
#* substitute goods or services; loss of use, data, or profits; or *
#* business interruption) however caused and on any theory of liability, *
#* whether in contract, strict liability, or tort (including negligence *
#* or otherwise) arising in any way out of the use of this software, even *
#* if advised of the possibility of such damage. *
#* *
#******************************************************************************
#
# To Do:
#
=head1 NAME
LTRPipeline - Run a structural LTR finder on the genome and polish the results
=head1 SYNOPSIS
LTRPipeline [-version] [-threads #] <input_sequence.fa>
=head1 DESCRIPTION
The options are:
=over 4
=item -version
Displays the version of the program
=back
=head1 CONFIGURATION OVERRIDES
=head1 SEE ALSO
RepeatModeler
=head1 COPYRIGHT
Copyright 2019-2022 Institute for Systems Biology
=head1 LICENSE
This code may be used in accordance with the Open Source License v. 3.0
http://opensource.org/licenses/OSL-3.0
=head1 AUTHORS
Robert Hubley <rhubley@systemsbiology.org>
Jullien Michelle Flynn <jmf422@cornell.edu>
=cut
#
# Module Dependence
#
use strict;
use Getopt::Long;
use Data::Dumper;
use Pod::Text;
use File::Path;
use File::Basename;
use FindBin;
use Cwd;
use Time::HiRes qw( gettimeofday tv_interval);
use lib $FindBin::RealBin;
use RepModelConfig;
use SeedAlignment;
use SeedAlignmentCollection;
#
# Magic numbers/constants here
# ie. my $PI = 3.14159;
#
my $DEBUG = 0;
$DEBUG = 1 if ( $RepModelConfig::DEBUGALL == 1 );
$| = 1; # Turn autoflush on
#### DEVELOPMENT ONLY #####
my $useLtrDet = 0;
#
# Version
#
my $version = $RepModelConfig::VERSION;
#
# Option processing
# e.g.
# -t: Single letter binary option
# -t=s: String parameters
# -t=i: Number paramters
#
my @getopt_args = (
'-version', # print out the version and exit
'-tmpdir=s',
'-debug',
'-LTRSeqLimit=i',
'-giToID=s',
'-LTRMaxSeqLen=i',
'-threads=i',
);
# Add configuration parameters as additional command-line options
push @getopt_args, RepModelConfig::getCommandLineOptions();
my %options = ();
Getopt::Long::config( "noignorecase", "bundling_override" );
unless ( GetOptions( \%options, @getopt_args ) ) {
usage();
}
$DEBUG = 1 if ( exists $options{'debug'} );
#
# Provide the POD text from this file and
# from the config file by merging them
# together. The heading "CONFIGURATION
# OVERRIDES" provides the insertion point
# for the configuration POD.
#
sub usage {
my $p = Pod::Text->new();
$p->output_fh( *STDOUT );
my $pod_str;
open IN, "<$0"
or die "Could not open self ($0) for generating documentation!";
while ( <IN> ) {
if ( /^=head1\s+CONFIGURATION OVERRIDES\s*$/ ) {
my $c_pod = RepModelConfig::getPOD();
if ( $c_pod ) {
$pod_str .= $_ . $c_pod;
}
}
else {
$pod_str .= $_;
}
}
close IN;
print "$0 - $version\n";
$p->parse_string_document( $pod_str );
exit( 1 );
}
if ( $options{'version'} ) {
print "$version\n";
exit;
}
#
# Resolve configuration settings using the following precedence:
# command line first, then environment, followed by config
# file.
#
RepModelConfig::resolveConfiguration( \%options );
my $config = $RepModelConfig::configuration;
#my $LTR_DETECTOR_DIR = $config->{'LTR_DETECTOR_DIR'}->{'value'};
my $LTR_DETECTOR_DIR;
my $LTR_RETRIEVER_DIR = $config->{'LTR_RETRIEVER_DIR'}->{'value'};
my $MAFFT_DIR = $config->{'MAFFT_DIR'}->{'value'};
my $NINJA_DIR = $config->{'NINJA_DIR'}->{'value'};
my $GENOMETOOLS_DIR = $config->{'GENOMETOOLS_DIR'}->{'value'};
my $REPEATMASKER_DIR = $config->{'REPEATMASKER_DIR'}->{'value'};
my $RMBLAST_DIR = $config->{'RMBLAST_DIR'}->{'value'};
my $CDHIT_DIR = $config->{'CDHIT_DIR'}->{'value'};
my $TRF_DIR = $config->{'TRF_DIR'}->{'value'};
my $UCSCTOOLS_DIR = $config->{'UCSCTOOLS_DIR'}->{'value'};
my $ltrSeqLimit = 10000;
$ltrSeqLimit = $options{'LTRSeqLimit'} if ( $options{'LTRSeqLimit'} );
#
# Run LTRDetector
# Run LTR_retreiver
# - runs RepeatMasker
# - runs cd-hit
# Run MAFFT
# Run NINJA
# Produce seed alignments and consensi
#
my $threads = 1;
if ( $options{'threads'} ) {
$threads = $options{'threads'};
}
unless ( -s $ARGV[ 0 ] ) {
usage();
}
my $inputFile = $ARGV[ 0 ];
## Temp directory
my $tempDir;
if ( $options{'tmpdir'} ) {
$tempDir = &createTempDir( [ $options{'tmpdir'}, cwd() ], "LTR" );
}
else {
$tempDir = &createTempDir( [ dirname( $inputFile ), cwd() ], "LTR" );
}
my %TimeBefore = ();
elapsedTime( "runtime" ); # Setup the timers
# Run LTR Detector
elapsedTime( "step" );
if ( $useLtrDet ) {
print "Running LtrDetector...";
my $numAnnots =
&runLtrDetector( $LTR_DETECTOR_DIR, $inputFile,
"$tempDir/raw-struct-results.txt",
$threads, $tempDir );
if ( $numAnnots == 0 ) {
print
"LTRPipeline: No results returned from LTR structural finder ( LtrDetector ).\n";
exit;
}
}
else {
print "Running LtrHarvest...";
my $numAnnots =
&runLtrHarvest( $GENOMETOOLS_DIR, $inputFile,
"$tempDir/raw-struct-results.txt",
$threads, $tempDir );
if ( $numAnnots == 0 ) {
print
"LTRPipeline: No results returned from LTR structural finder ( LtrHarvest ).\n";
exit;
}
}
print " : " . elapsedTime( "step" ) . "\n";
# Postprocess results using LTR_Retriever
elapsedTime( "step" );
print "Running Ltr_retriever...";
&runLtrRetriever(
$LTR_RETRIEVER_DIR,
$inputFile,
"$tempDir/raw-struct-results.txt",
"$tempDir/LtrRetriever-redundant-results.fa",
$threads,
$ltrSeqLimit
);
if ( !-s "$tempDir/LtrRetriever-redundant-results.fa" ) {
print "LTRPipeline: No results after LTR_Retriever filtering.\n";
exit;
}
print " : " . elapsedTime( "step" ) . "\n";
#
# Filter out sequences greater than LTRMaxSeqLen if requested
#
if ( $options{'LTRMaxSeqLen'} && $options{'LTRMaxSeqLen'} =~ /\d+/ ) {
system(
"mv $tempDir/LtrRetriever-redundant-results.fa $tempDir/LtrRetriever-redundant-results-orig.fa"
);
open IN, "<$tempDir/LtrRetriever-redundant-results-orig.fa"
or die
"Could not open $tempDir/LtrRetriever-redundant-results-orig.fa for reading!\n";
open OUT, ">$tempDir/LtrRetriever-redundant-results.fa"
or die
"Could not open $tempDir/LtrRetriever-redundant-results.fa for writing!\n";
my $seq = "";
my $id = "";
while ( <IN> ) {
if ( /^>(\S+)/ ) {
my $tmpID = $1;
if ( $seq ) {
if ( length( $seq ) <= $options{'LTRMaxSeqLen'} ) {
print OUT ">$id\n$seq\n";
}
}
$seq = "";
$id = $tmpID;
next;
}
s/[\n\r\s]+//g;
$seq .= $_;
}
if ( $seq ) {
if ( length( $seq ) <= $options{'LTRMaxSeqLen'} ) {
print OUT ">$id\n$seq\n";
}
}
close IN;
close OUT;
}
# Group elements into families using a scoring system that accounts for common deletion products
elapsedTime( "step" );
print "Aligning instances...";
&runMafft( $MAFFT_DIR, "$tempDir/LtrRetriever-redundant-results.fa",
"$tempDir/mafft-alignment.fa", $threads );
if ( !-s "$tempDir/mafft-alignment.fa" ) {
print
"LTRPipeline: Error - could not produce a multiple alignment from the denovo LTR results.\n";
exit;
}
print " : " . elapsedTime( "step" ) . "\n";
#
elapsedTime( "step" );
print "Clustering...";
&runNinja( $NINJA_DIR, "$tempDir/mafft-alignment.fa", "$tempDir/clusters.dat",
$threads );
if ( !-s "$tempDir/clusters.dat" ) {
print "LTRPipeline: Error - could not cluster MAFFT results.\n";
}
print " : " . elapsedTime( "step" ) . "\n";
#
# Here Jullien screened out families which have similarity to DNA Transposons or LINE elements by searching
# with RepeatMasker. Since this is a de-novo approach we do not expect there to be a library available.
# One could search against all known LINE/DNA elements but that will need further thought.
#
# Read in cluster data
open IN, "<$tempDir/clusters.dat"
or die "LTRPipeline : Error - could not open $tempDir/clusters.dat!";
my %clusters;
while ( <IN> ) {
# 0 utg7180_2262700..2263120_LTR#LTR/Gypsy
if ( /^(\d+)\s+(\S+)\s*$/ ) {
push @{ $clusters{$1} }, $2;
}
}
close IN;
# Read in Ltr_Retriever sequences
open IN, "<$tempDir/LtrRetriever-redundant-results.fa"
or die
"LTRPipeline : Error - could not open $tempDir/LtrRetriever-redundant-results.fa!";
my %seeds = ();
my $seq = "";
my $id = "";
while ( <IN> ) {
if ( /^>(\S+)/ ) {
my $tmpID = $1;
if ( $seq ) {
$seeds{$id} = $seq;
}
$seq = "";
$id = $tmpID;
next;
}
s/[\n\r\s]+//g;
$seq .= $_;
}
if ( $seq ) {
$seeds{$id} = $seq;
}
close IN;
# Process each cluster
elapsedTime( "step" );
print "Refining families...";
open OUTSTK, ">$tempDir/families.stk"
or die "Could not create $tempDir/families.stk!\n";
open OUTCON, ">$tempDir/families.fa"
or die "Could not create $tempDir/families.fa!\n";
foreach my $cluster ( sort keys %clusters ) {
my $idx = $cluster + 1;
# Make a fasta file for family ( cluster ).
open OUT, ">$tempDir/ltr-1_family-$idx.fa"
or die "LTRPipeline : Could not create $tempDir/ltr-1_family-$idx.fa!";
my $intCount = 0;
my $ltrCount = 0;
foreach my $seedID ( @{ $clusters{$cluster} } ) {
if ( $seedID =~ /_LTR/ ) {
$ltrCount++;
}
else { $intCount++; }
print OUT ">$seedID\n$seeds{$seedID}\n";
}
close OUT;
my $famType = "LTR";
$famType = "INT" if ( $intCount > $ltrCount );
# Run Refiner on the seeds
my $additionalOpts = "";
$additionalOpts .= "-giToID " . $options{'giToID'} if ( $options{'giToID'} );
my $cmd =
"$FindBin::RealBin/Refiner -noTmp $additionalOpts -name ltr-1_family-$idx $tempDir/ltr-1_family-$idx.fa";
$cmd .= " -quiet" unless ( $DEBUG );
print "LTRPipeline: Running $cmd\n" if ( $DEBUG );
system( $cmd );
# family-72.fa.refiner.stk
if ( -s "$tempDir/ltr-1_family-$idx.fa.refiner_cons" ) {
my $cons;
my $maSize;
open INREF, "<$tempDir/ltr-1_family-$idx.fa.refiner_cons"
or die "LTRPipeline"
. ": Could not open refined model $tempDir/ltr-1_family-$idx.fa.refiner_cons!\n";
while ( <INREF> ) {
if ( /Final Multiple Alignment Size = (\d+)/ ) {
$maSize = $1;
}
else {
$cons .= $_;
}
}
close INREF;
# Clean up consensi starting/ending with Ns
# - These are sections of the multiple alignment
# where there is no support for a consensus.
$cons =~ s/^[Nn]*([^Nn].*[^Nn])[Nn]*$/$1/;
if ( -s "$tempDir/ltr-1_family-$idx.fa.refiner.stk" ) {
my $stockholmFile = SeedAlignmentCollection->new();
my $IN;
open $IN, "<$tempDir/ltr-1_family-$idx.fa.refiner.stk"
or die
"Could not open $tempDir/ltr-1_family-$idx.fa.refiner.stk for reading!\n";
$stockholmFile->read_stockholm( $IN );
close $IN;
# File only contains one family
my $seedAlign = $stockholmFile->get( 0 );
# Add details to DE line
$seedAlign->setDescription( "RepeatModeler Generated - ltr-1"
. "_family-$idx, [ Type=$famType, Final Multiple Alignment Size = $maSize ]"
);
print OUTSTK "" . $seedAlign->toString();
}
# Save the consensus to the consensi file.
print OUTCON ">ltr-1_family-$idx [ Type=$famType, "
. "Final Multiple Alignment Size = "
. $maSize . " ]\n";
print OUTCON "$cons\n";
}
}
print " : " . elapsedTime( "step" ) . "\n";
close OUTCON;
close OUTSTK;
system( "cp $tempDir/families.fa $inputFile-ltrs.fa" )
if ( -s "$tempDir/families.fa" );
system( "cp $tempDir/families.stk $inputFile-ltrs.stk" )
if ( -s "$tempDir/families.stk" );
print "Program Time: " . elapsedTime( "runtime" ) . "\n";
exit;
###############################################################################################
sub runNinja {
my $binDir = shift;
my $alnFile = shift;
my $outFile = shift;
my $threads = shift;
my $wrkDir = shift;
die "LTRPipeline::runNinja : Could not find sequence input file $alnFile\n"
if ( !-s $alnFile );
# Create temporary directory
my $tmpDir;
if ( !defined $wrkDir || !-d $wrkDir ) {
$tmpDir = &createTempDir( [ dirname( $alnFile ), cwd() ], "NINJA" );
}
else {
$tmpDir = &createTempDir( [ $wrkDir ], "NINJA" );
}
print "LTRPipeline::runNinja : tmpdir = $tmpDir\n" if ( $DEBUG );
my $cmd =
"$binDir/Ninja --in $alnFile --out $tmpDir/cluster.dat --out_type c --corr_type m --cluster_cutoff 0.2";
if ( defined $threads && $threads > 1 ) {
$cmd .= " --threads $threads";
}
$cmd .= " > $tmpDir/Ninja.log 2>&1";
print "LTRPipeline::runNinja : Running analysis $cmd\n" if ( $DEBUG );
system( $cmd);
if ( -s "$tmpDir/cluster.dat" ) {
system( "mv $tmpDir/cluster.dat $outFile" );
}
# TODO: Consider keeping on error
unless ( $DEBUG ) {
rmtree( $tmpDir );
}
}
sub runMafft {
my $binDir = shift;
my $seqFile = shift;
my $outFile = shift;
my $threads = shift;
my $wrkDir = shift;
die "LTRPipeline::runMafft : Could not find sequence input file $seqFile\n"
if ( !-s $seqFile );
# Create temporary directory
my $tmpDir;
if ( !defined $wrkDir || !-d $wrkDir ) {
$tmpDir = &createTempDir( [ dirname( $seqFile ), cwd() ], "MAFFT" );
}
else {
$tmpDir = &createTempDir( [ $wrkDir ], "MAFFT" );
}
$ENV{MAFFT_TMPDIR} = $tmpDir;
print "LTRPipeline::runMafft : tmpdir = $tmpDir\n" if ( $DEBUG );
# NOTE: --quiet hides some error reporting. Need to develop a way
# to catch common errors like:
#
# nthread = 20
# nthreadpair = 20
# nthreadtb = 16
# ppenalty_ex = 0
# stacksize: 8192 kb
# generating a scoring matrix for nucleotide (dist=200) ... done
# Gap Penalty = -1.53, +0.00, +0.00
# =========================================================================
# =========================================================================
# ===
# === Alphabet 'i' is unknown.
# === Please check site 8234 in sequence 2.
# ===
# === To make an alignment that has unusual characters (U, @, #, etc), try
# === % mafft --anysymbol input > output
# ===
# =========================================================================
# =========================================================================
# Illegal character i
#
my $cmd = "$binDir/mafft --large --quiet";
if ( defined $threads && $threads > 1 ) {
$cmd .= " --thread $threads";
}
$cmd .= " $seqFile > $tmpDir/mafft-alignment.fa";
print "LTRPipeline::runMafft : Running analysis $cmd\n" if ( $DEBUG );
system( $cmd);
if ( -s "$tmpDir/mafft-alignment.fa" ) {
system( "mv $tmpDir/mafft-alignment.fa $outFile" );
}
# TODO: Consider keeping on error
unless ( $DEBUG ) {
rmtree( $tmpDir );
}
}
sub runLtrRetriever {
my $binDir = shift;
my $seqFile = shift;
my $annotFile = shift;
my $outFile = shift;
my $threads = shift;
my $ltrSeqLimit = shift;
my $wrkDir = shift;
die
"LTRPipeline::runLtrRetriever : Could not find sequence input file $seqFile\n"
if ( !-s $seqFile );
die
"LTRPipeline::runLtrRetriever : Could not find annotation input file $annotFile\n"
if ( !-s $annotFile );
# Create temporary directory
my $tmpDir;
if ( !defined $wrkDir || !-d $wrkDir ) {
$tmpDir = &createTempDir( [ dirname( $annotFile ), cwd() ], "LRET" );
}
else {
$tmpDir = &createTempDir( [ $wrkDir ], "LRET" );
}
print "LTRPipeline::runLtrRetriever : tmpdir = $tmpDir\n" if ( $DEBUG );
# Create a symlink inside the temp directory called "seq.fa" pointing
# to the sequence file ( using full path ). LTR_retriever creates
# a prodigous amount of temporary files and output files in the same
# directory as the sequence file. This ensures they all get created
# inside the temporary directory.
my $full_seqFile = File::Spec->rel2abs( $seqFile );
symlink( $full_seqFile, "$tmpDir/seq.fa" );
my $full_annotFile = File::Spec->rel2abs( $annotFile );
# NOTE: in LTR_retriever 2.9.0 trf_path is not the path
# to the directory containing the binary but rather
# the full path to the binary itself.
my $cmd = "cd $tmpDir; $binDir/LTR_retriever " .
" -repeatmasker $REPEATMASKER_DIR" .
" -blastplus $RMBLAST_DIR" .
" -cdhit_path $CDHIT_DIR" .
" -trf_path $TRF_DIR/trf" .
" -genome seq.fa -inharvest $full_annotFile -noanno";
if ( $threads > 1 ) {
$cmd .= " -threads $threads";
}
$cmd .= " > LTR_retriever.log 2>&1";
print "LTRPipeline::runLtrRetriever : Running analysis $cmd\n" if ( $DEBUG );
my $cwd = cwd();
system( $cmd);
chdir $cwd
or die
"LTRPipeline::runLtrRetriever : Could not change directory back to $cwd : $!";
my $redundantLibFile = "";
my $error = 0;
if ( -s "$tmpDir/seq.fa.LTRlib.redundant.fa" ) {
$redundantLibFile = "$tmpDir/seq.fa.LTRlib.redundant.fa";
}
elsif ( -s "$tmpDir/seq.fa.mod.LTRlib.redundant.fa" ) {
$redundantLibFile = "$tmpDir/seq.fa.mod.LTRlib.redundant.fa";
}
else {
#print "\nERROR: LTRPipeline::runLtrRetriever : Could not locate redundant library in $tmpDir\n";
$error = 1;
}
my $count = 0;
if ( $redundantLibFile ) {
open IN, "<$redundantLibFile"
or die
"LTRPipeline::runLtrRetriever : Could not open $redundantLibFile for reading!\n";
my $numRedundant = 0;
while ( <IN> ) {
$numRedundant++ if ( /^>/ );
}
close IN;
if ( $numRedundant > $ltrSeqLimit ) {
my $nonRedundantLibFile;
if ( -s "$tmpDir/seq.fa.mod.LTRlib.fa" ) {
$nonRedundantLibFile = "$tmpDir/seq.fa.mod.LTRlib.fa";
}
elsif ( -s "$tmpDir/seq.fa.LTRlib.fa" ) {
$nonRedundantLibFile = "$tmpDir/seq.fa.LTRlib.fa";
}
if ( $nonRedundantLibFile ) {
open IN, "<$nonRedundantLibFile"
or die
"LTRPipeline::runLtrRetriever : Could not open $nonRedundantLibFile for reading!\n";
my $numNonRedundant = 0;
while ( <IN> ) {
$numNonRedundant++ if ( /^>/ );
}
close IN;
if ( $numNonRedundant < $ltrSeqLimit ) {
print
"INFO: LTRPipeline - redundant library too large ( $numRedundant ). Falling back to non-redundant library ( $numNonRedundant ).\n";
system( "mv $nonRedundantLibFile $outFile" );
$count = $numRedundant;
}
else {
print
"ERROR: LTRPipeline::runLtrRetriever : LTR libraries ( redundant: $numRedundant, non-redundant: $numNonRedundant ) are larger than permissible threshold $ltrSeqLimit ). See $tmpDir for details.\n";
$error = 1;
}
}
else {
print
"ERROR: LTRPipeline::runLtrRetriever : Could not find non-redundant library $tmpDir/seq.fa.mod.LTRlib.fa or $tmpDir/seq.fa.LTRlib.fa!\n";
$error = 1;
}
}
elsif ( $numRedundant > 0 ) {
system( "mv $redundantLibFile $outFile" );
$count = $numRedundant;
}
}
# Currently there isn't a way to get the orientation of the sequences other than comparing them with
# the input. So here we go:
# Read in the library sequences and ranges
open IN,"<$outFile" or die "LTR_retriever failed to generate a file. Please check $tmpDir/LTR_retriever.log for details.";
my $seqID = "";
my $start = 0;
my $end = 0;
my $seq = "";
my $suffix = "";
my @lib = ();
while ( <IN> ) {
# e.g. >utg7180:3885179..3890145_INT#LTR/Gypsy
if ( /^>(\S+):(\d+)\.\.(\d+)(.*)/ ) {
my $tmpSeqID = $1;
my $tmpStart = $2;
my $tmpEnd = $3;
my $tmpSuffix = $4;
if ( $seqID ne "" ) {
push @lib,[ $seqID, $start, $end, $seq, $suffix];
}
$seqID = $tmpSeqID;
$start = $tmpStart;
$end = $tmpEnd;
$seq = "";
$suffix = $tmpSuffix;
next;
}
s/[\n\r\s]+//g;
$seq .= $_;
}
if ($seqID ne ""){
push @lib,[ $seqID, $start, $end, $seq, $suffix];
}
close IN;
# Using the sequence file determine the orientation of each sequence
# NOTE: This also validates the sequence ranges and flags LTR_retriever
# results that have incorrect ranges
getOrientation(\@lib, $seqFile);
# Backup original outFile and regenerate with oriented ranges.
system("mv $outFile $outFile.no_orient");
open OUT,">$outFile" or die;
foreach my $entry ( @lib ) {
my $type = "";
if ( $entry->[4] =~ /_LTR/ ) {
$type = "_LTR";
}elsif ( $entry->[4] =~ /_INT/ ) {
$type = "_INT";
}
my $id = $entry->[0] . ":";
if ( $entry->[5] eq "!" ) {
$id .= $entry->[1] . ".." . $entry->[2] . "$type [warning: bad ltr_retriever range]";
}elsif ( $entry->[5] eq "+" ) {
$id .= $entry->[1] . ".." . $entry->[2] . "$type [forward]";
}else {
$id .= $entry->[2] . ".." . $entry->[1] . "$type [reverse]";
}
print OUT ">$id\n$entry->[3]\n";
}
close OUT;
unless ( $DEBUG || $error ) {
rmtree( $tmpDir );
}
return $count;
}
sub getOrientation {
my $lib = shift;
my $seqFile = shift;
system("$UCSCTOOLS_DIR/faToTwoBit -long $seqFile $seqFile.2bit");
if ( ! -s "$seqFile.2bit" ) {
die "getOrientation: Something went wrong calling $UCSCTOOLS_DIR/faToTwoBit on $seqFile!\n";
}
my %lookup = ();
open OUT,">$seqFile.seqlist" or die;
foreach my $entry ( @{$lib} ) {
my $twoBitID = "$entry->[0]:" . ($entry->[1]-1) . "-$entry->[2]";
print OUT "$twoBitID\n";
$lookup{$twoBitID} = $entry;
}
close OUT;
open IN,"$UCSCTOOLS_DIR/twoBitToFa -seqList=$seqFile.seqlist $seqFile.2bit stdout|" or
die "getOrientation: Something went wrong calling $UCSCTOOLS_DIR/twoBitToFa on $seqFile.2bit!\n";
my $resID = "";
my $seq;
while ( <IN> ) {
if ( /^>(\S+)/ ) {
my $tID = $1;
if ( $seq ne "" )
{
if ( exists $lookup{$resID} )
{
if ( $seq eq $lookup{$resID}->[3] ) {
# Forward range
push @{$lookup{$resID}}, "+";
}else {
$seq = reverse($seq);
$seq =~ tr/ACGTN/TGCAN/;
if ( $seq eq $lookup{$resID}->[3] ) {
# Reverse range
push @{$lookup{$resID}}, "-";
}else {
# Invalid range
push @{$lookup{$resID}}, "!";
}
}
}else {
warn "Oops...couldn't locate match to $resID in lookup table!\n";
}
}
$resID = $tID;
$seq = "";
next;
}
s/[\n\r\s]+//g;
$seq .= $_;
}
if ( $seq ne "" )
{
if ( exists $lookup{$resID} )
{
if ( $seq eq $lookup{$resID}->[3] ) {
# Forward range
push @{$lookup{$resID}}, "+";
}else {
$seq = reverse($seq);
$seq =~ tr/ACGTN/TGCAN/;
if ( $seq eq $lookup{$resID}->[3] ) {
# Reverse range
push @{$lookup{$resID}}, "-";
}else {
# Invalid range
push @{$lookup{$resID}}, "!";
}
}
}else {
warn "getOrientation: Oops...couldn't locate match to $resID in lookup table!\n";
}
}
close IN;
}
sub runLtrHarvest {
my $binDir = shift;
my $seqFile = shift;
my $outFile = shift;
my $threads = shift;
my $wrkDir = shift;
die "LTRPipeline::runLtrHarvest : Could not find input file $seqFile\n"
if ( !-s $seqFile );
# Flag to maintain files should an error occur
my $errorFlag = 0;
# Total number of annotations found
my $annotCnt = 0;
# Create temporary directory
my $tmpDir;
if ( !defined $wrkDir || !-d $wrkDir ) {
$tmpDir = &createTempDir( [ dirname( $seqFile ), cwd() ], "LHAR" );
}
else {
$tmpDir = &createTempDir( [ $wrkDir ], "LHAR" );
}
print "LTRPipeline::runLtrHarvest : tmpdir = $tmpDir\n" if ( $DEBUG );
# Build enhanced suffix array datastructure for Ltr_harvest
# Must run in the tmpDir as this program will insist on creating
# esa_* files in the working directory only
my $full_seqFile = File::Spec->rel2abs( $seqFile );
my $origDir = getcwd();
my $addlOpts = "";
# There is a bug in 1.5.9 and 1.5.10 that appears to not generate the correct files
# if -j is used.
#$addlOpts = "-j $options{'threads'} " if ( $options{'threads'} );
chdir( $tmpDir );
my $cmd =
"$binDir/gt $addlOpts suffixerator -db $full_seqFile -indexname esa_index -tis -suf -lcp -des -ssp -sds -dna > suffixerator.log 2>&1";
my $retCode = system( $cmd);
if ( $retCode != 0 ) {
warn
"LtrPipeline: GenomeTools failed to build suffixtree datastructure. This is frequently\n"
. " caused by a lack of disk space. Error code: "
. ( $? >> 8 ) . "\n";
$errorFlag = 1;
chdir( $origDir );
}
else {
# Run ltrharvest
$cmd =
"$binDir/gt $addlOpts ltrharvest -index esa_index -out ltrharvest.out > ltrharvest.log 2>&1";
$retCode = system( $cmd);
chdir( $origDir );
if ( $retCode != 0 ) {
warn "LtrPipeline: GenomeTools failed to run ltrharvest. Error code: "
. ( $? >> 8 ) . " Run on esa-index created from: $full_seqFile\n";
$errorFlag = 1;
}
if ( -s "$tmpDir/ltrharvest.log" ) {
open IN, "<$tmpDir/ltrharvest.log"
or die "Could not open $tmpDir/ltrharvest.log for reading!\n";
open OUT, ">$outFile" or die "Could not open $outFile for writing!\n";
while ( <IN> ) {
if ( /^#/ ) {
print OUT;
}
elsif ( /^\d+\s+\d+\s+\d+\s+\d+/ ) {
$annotCnt++;
print OUT;
}
elsif ( /^\s*$/ ) {
# Don't save
}
else {
warn
"LtrPipeline: Ltrharvest returned an unexpected result line:\n$_";
$errorFlag = 1;
}
}
close IN;
close OUT;
}
else {
# Expected some sort of output
$errorFlag = 1;
}
}
unless ( $DEBUG || $errorFlag ) {
rmtree( $tmpDir );
}
print "LTRPipeline::runLtrHarvest : Returning $annotCnt annotations.\n"
if ( $DEBUG );
return $annotCnt;
}
sub runLtrDetector {
my $binDir = shift;
my $seqFile = shift;
my $outFile = shift;
my $threads = shift;
my $wrkDir = shift;
die "LTRPipeline::runLtrDetector : Could not find input file $seqFile\n"
if ( !-s $seqFile );
# The initial release of LtrDetector appears to only want one
# sequence per input file.
open INN, "<$seqFile"
or die
"LTRPipeline::runLtrDetector : Could not find open input file $seqFile\n";
# Create temporary directory
my $tmpDir;
if ( !defined $wrkDir || !-d $wrkDir ) {
$tmpDir = &createTempDir( [ dirname( $seqFile ), cwd() ], "LDET" );
}
else {
$tmpDir = &createTempDir( [ $wrkDir ], "LDET" );
}
print "LTRPipeline::runLtrDetector : tmpdir = $tmpDir\n" if ( $DEBUG );
my $id;
my $seq;
my $i = 1;
while ( <INN> ) {
if ( /^>(\S+)/ ) {
my $tmpID = $1;
if ( $seq ) {
# Create temporary files
open OUT, ">$tmpDir/ltrdetector-batch-$i.fa"
or die
"LTRPipeline::runLtrDetector : Could not create $tmpDir/ltrdetector-batch-$i.fa\n";
print OUT ">$id\n$seq\n";
close OUT;
$i++;
}
$id = $tmpID;
$seq = "";
next;
}
s/[\n\r\s]+//g;
$seq .= $_;
}
close INN;
if ( $seq ) {
# Create temporary files
open OUT, ">$tmpDir/ltrdetector-batch-$i.fa"
or die
"LTRPipeline::runLtrDetector : Could not create $tmpDir/ltrdetector-batch-$i.fa\n";
print OUT ">$id\n$seq\n";
close OUT;
$i++;
}