annotateM

#!/usr/bin/env perl
###############################################################################
#
#    annotateM
#    
#    The idea here is an automated way of annotating your genome based on 
#    multiple available databases and to produce a tab-delimited file of 
#    all the annotations, evalues, scores, descriptions.
# 
#    Suggested workflow:
#	 1) run your genome nucleotide fasta file through annotateM
#	 2) generate a tab-delimited file
#	 3) open the file in ms excel or oo calc
#	 4) manually curate the annotations based on evalues/scores/desc etc	
#        5) metabolic reconstruction of organism
#
#    Copyright (C) Mohamed Fauzi Haroon
#	 Special appearance from Adam Skarshewski
#
#    This program is free software: you can redistribute it and/or modify
#    it under the terms of the GNU General Public License as published by
#    the Free Software Foundation, either version 3 of the License, or
#    (at your option) any later version.
#
#    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.
#
#    You should have received a copy of the GNU General Public License
#    along with this program.  If not, see <http://www.gnu.org/licenses/>.
#
###############################################################################

#pragmas
use strict;
use warnings;

#core Perl modules
use Getopt::Long;
use Carp;
use Data::Dumper;

#CPAN modules

#locally-written modules

BEGIN {
    select(STDERR);
    $| = 1;
    select(STDOUT);
    $| = 1;
}

# edit here to log all external commands 
my $global_log_commands = 0;

# ext command failure levels
use constant {
    IGNORE_FAILURE => 0,
    WARN_ON_FAILURE => 1,
    DIE_ON_FAILURE => 2
};

# get input params and print copyright
printAtStart();
my $global_options = checkParams();

######################################################################
# CODE HERE
######################################################################

# check that the file exists
checkFileExists($global_options->{'in'});

if (! -e "./prokka_annotation/")
{
#print "prokka\n";
# run prokka to generate the ORFs and also prokka annotations
checkAndRunCommand("prokka", [{
                                   "--locustag" =>  $global_options->{'locustag'},
				   "--outdir" => "prokka_annotation",
				   "--prefix" => $global_options->{'locustag'},
				   "--kingdom" =>  $global_options->{'kingdom'},
				   "--cpus" =>  $global_options->{'threads'},
				   "--keep_names",
				   $global_options->{'in'},	
                                   }], DIE_ON_FAILURE);
}

# identify the ORF called amino acid fasta file for blast-ing
my $locus = $global_options->{'locustag'};

# blast against img
if (! -e "./$locus.faaVSimg.blastp")
{
print "BLASTing against IMG 4.0 database...............\n";
checkAndRunCommand("cat", 
			[[ 
			"prokka_annotation/$locus.faa |",	
			"parallel",
			"--block"=> "100k", 
			"--recstart",
			"'>'",
			"--pipe",
			"blastp",
			-db => "/srv/db/img/4.0/dereplicated/img_dereplicated_species.genes.faa",
			-outfmt => 6,
			-max_target_seqs => 1,
			-evalue => $global_options->{'evalue'},
			-query => "-",
			"> $locus.faaVSimg.blastp",
			]], DIE_ON_FAILURE);
}

# reciprocal blast of img positive hits against genome ORF
if (! -e "./subsetimg.faaVS$locus.faa.blastp")
{
print "Reciprocal BLASTing positive IMG hits to $locus.faa ...............\n";
checkAndRunCommand("contig_extractor.pl",
			[[
			-i => "$locus.faaVSimg.blastp",
			-d => "/srv/db/img/4.0/dereplicated/img_dereplicated_species.genes.faa",
			-b => '',
			-S => '',
			-o => "subsetimg.faa",
			]], DIE_ON_FAILURE);

checkAndRunCommand("makeblastdb",
			[[
			-in => "prokka_annotation/$locus.faa",
			-dbtype => "prot",
			]], DIE_ON_FAILURE);

checkAndRunCommand("blastp",
			[[
			-query => "subsetimg.faa",
			-db => "prokka_annotation/$locus.faa",
			-outfmt => 6,
			-max_target_seqs => 1,
			-evalue => $global_options->{'evalue'},
			-num_threads => $global_options->{'threads'},
			-out => "subsetimg.faaVS$locus.faa.blastp",
			]], DIE_ON_FAILURE);
}

# blast against uniref				
if (! -e "./$locus.faaVSuniref90.blastp")
{
print "BLASTing against Uniref90 database................\n";
checkAndRunCommand("cat",[[
			"prokka_annotation/$locus.faa |",
                        "parallel",
                        "--block"=> "100k",
                        "--recstart",
                        "'>'",
                        "--pipe",
			"blastp",
			-db => "/srv/db/uniprot/uniref-20140403/uniref90.fasta",
			-outfmt => 6,
                        -max_target_seqs => 1,
			-evalue => $global_options->{'evalue'},
                        -query => "-",
                        "> $locus.faaVSuniref90.blastp",
                        #-num_threads => $global_options->{'threads'},
                        ]], DIE_ON_FAILURE);
}

# reciprocal blast of Uniref positive hits against genome ORF
if (! -e "./subsetuniref.faaVS$locus.faa.blastp")
{
print "Reciprocal BLASTing positive Uniref hits to $locus.faa ...............\n";

checkAndRunCommand("contig_extractor.pl",
                        [[
                        -i => "$locus.faaVSuniref90.blastp",
                        -d => "/srv/db/uniprot/uniref-20140403/uniref90.fasta",
                        -b => '',
                        -S => '',
                        -o => "subsetuniref.faa",
                        ]], DIE_ON_FAILURE);

checkAndRunCommand("blastp",
                        [[
                        -query => "subsetuniref.faa",
                        -db => "prokka_annotation/$locus.faa",
                        -outfmt => 6,
                        -max_target_seqs => 1,
                        -evalue => $global_options->{'evalue'},
                        -num_threads => $global_options->{'threads'},
                        -out => "subsetuniref.faaVS$locus.faa.blastp",
                        ]], DIE_ON_FAILURE);
}

# blast against COG
if (! -e "./$locus.faaVSCOG.blastp")
{
print "BLASTing against COG database................\n";
checkAndRunCommand("cat",[[
                        "prokka_annotation/$locus.faa |",
                        "parallel",
                        "--block"=> "100k",
                        "--recstart",
                        "'>'",
                        "--pipe",
                        "blastp",
                        -db => "/srv/db/cog/cog_blast_prot_db",
                        -outfmt => 6,
                        -max_target_seqs => 1,
                        -evalue => $global_options->{'evalue'},
                        -query => "-",
                        "> $locus.faaVSCOG.blastp",
                        #-num_threads => $global_options->{'threads'},
                         ]], DIE_ON_FAILURE);
}

# HMMSCAN against PFAM
if (! -e "./$locus.faaVSPfam-A.hmm.hmmscanned")
{
print "HMMscanning against Pfam database................\n";
checkAndRunCommand("pfam_scan.pl",[[
			-cpu => $global_options->{'threads'}, 
			-e_seq => $global_options->{'evalue'},
			-outfile => "$locus.faaVSPfam-A.hmm.hmmscanned",
			-fasta => "prokka_annotation/$locus.faa",
			-dir => "/srv/db/pfam/27",
			]], DIE_ON_FAILURE);
}

# HMMSCAN against TIGRfam
if (! -e "./$locus.faaVStigr_all.hmm.hmmscanned")
{
print "HMMscanning against TIGRfam database................\n";
checkAndRunCommand("hmmscan",[[
			"--tblout",
			"$locus.faaVStigr_all.hmm.hmmscanned",
			"--noali",
			-E => $global_options->{'evalue'},
			"--cpu",
			 $global_options->{'threads'},
			"/srv/db/tigrfam/14.0/TIGRFAMs_14.0_HMM/tigr_all.hmm",
			"prokka_annotation/$locus.faa",
			]], DIE_ON_FAILURE);
}
			
# convert the hmmscan output to tab delimited
checkAndRunCommand("awk",[[
                       "'{\$1=\$1}{ print }'",
			"$locus.faaVSPfam-A.hmm.hmmscanned",
			"| sed 's/\\s/\\t/g'",
			"> $locus.faaVSPfam-A.hmm.hmmscanned.tab",
                      ]], DIE_ON_FAILURE);

checkAndRunCommand("awk",[[
			"'{\$1=\$1}{ print }'",
                       "$locus.faaVStigr_all.hmm.hmmscanned",
			"| sed 's/\\s/\\t/g'",
                       "> $locus.faaVStigr_all.hmm.hmmscanned.tab",
                       ]], DIE_ON_FAILURE);


# hashes for img
my %hash = ();
my %hash2 = ();
my %hash3 =();

# read the img blast output and store in hash
open my $IMGblast, "./$locus.faaVSimg.blastp", or die "Couldn't open file $locus.faaVSimg.blastp\n";
while (<$IMGblast>)
{
	chomp $_;
	my @columns = split (/\t/, $_);
	# store the orfid which is in columns[0]
	my $orfid = $columns[0];
	if ($columns[11] > 60)
	{
		# key is $orfid while the value of the key in $columns[1] is the IMG id 
		$hash{$orfid} = $columns[1];
		$hash{$columns[1]} = $columns[0]; 
		$hash2{$columns[1]} = "$columns[0]\t$columns[10]\t$columns[11]";	
	}
}

# read img id2names.txt
open my $imgid2names, "/srv/db/img/4.0/dereplicated/id2names.txt", or die "Couldn't open id2names.txt\n";
open my $img_temp_OUT, ">img_output_temp.txt";
while (<$imgid2names>)
{
	chomp $_;
	my @columns = split (/\t/, $_);
	if (exists $hash2{$columns[0]})
	{	
		$hash3{$columns[0]} = "$hash2{$columns[0]}\t$columns[1]\t$columns[2]";	
		print {$img_temp_OUT} "$hash2{$columns[0]}\t$columns[1]\t$columns[2]\n";
	}
}

close($IMGblast);
close($imgid2names);
close($img_temp_OUT);

# read my reciprocal img blast output and store in hash
open my $rIMGblast, "./subsetimg.faaVS$locus.faa.blastp", or die "Couldn't open file subsetimg.faaVS$locus.faa.blastp\n";
open my $img_temp_OUT2, ">img_output_temp2.txt";
while (<$rIMGblast>)
{
	chomp $_;
	my @columns = split (/\t/, $_);
	if (exists $hash{$columns[0]})
	{
		print {$img_temp_OUT2} "$hash3{$columns[0]}\treciprocal\n";
	}
}

close($img_temp_OUT2);

# hashes for uniref	
my %hash4 = ();
my %hash5 =();
my %hash6 = ();

# read uniref blast and store in hash
open my $unirefblast, "./$locus.faaVSuniref90.blastp", or die "Couldn't open file $locus.faaVSuniref90.blastp\n";
while (<$unirefblast>)
{
	chomp $_;
        my @columns = split (/\t/, $_);
	if ($columns[11] > 60)
	{
		$hash4{$columns[0]} = $columns[1];
		$hash4{$columns[1]} = $columns[0];
		$hash5{$columns[1]} = "$columns[0]\t$columns[10]\t$columns[11]";
	}
}

# read uniref id2names.txt
open my $unirefid2names, "/srv/db/uniprot/uniref-20140403/uniref90_id2names.txt", or die "Couldn't open id2names.txt\n";
open my $uniref_temp_OUT, ">uniref_output_temp.txt";
while (<$unirefid2names>)
{
	chomp $_;
        my @columns = split (/\t/, $_);
	if (exists $hash5{$columns[0]})
        {
                $hash6{$columns[0]} = "$hash5{$columns[0]}\t$columns[1]\t$columns[2]";
                print {$uniref_temp_OUT} "$hash5{$columns[0]}\t$columns[1]\t$columns[2]\n";
        }
}

close($unirefblast);
close($unirefid2names);
close($uniref_temp_OUT);

# read my reciprocal img blast output and store in hash
open my $runirefblast, "./subsetuniref.faaVS$locus.faa.blastp", or die "Couldn't open file subsetuniref.faaVS$locus.faa.blastp\n";
open my $uniref_temp_OUT2, ">uniref_output_temp2.txt";
while (<$runirefblast>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        if (exists $hash4{$columns[0]})
        {
            print {$uniref_temp_OUT2} "$hash6{$columns[0]}\treciprocal\n";
        }
}

close($uniref_temp_OUT2);

# hashes for pfam
my %hash7 = ();
my %hash8 = ();

# read pfam hmmscan output and store in hash
open my $pfamoutput, "./$locus.faaVSPfam-A.hmm.hmmscanned.tab", or die "Couldn't open file $locus.faaVSPfam-A.hmm.hmmscanned.tab\n";
while (<$pfamoutput>)
{
	next if /^\s*(#.*)?$/;
	next if $pfamoutput =~ /^#/;
	next if $pfamoutput =~ /^=/;
    chomp $_;
    my @columns = split (/\t/, $_);
    if ($columns[11] > 60)
    {
		my @pfam_columns = split (/\./, $columns[5]);
		my $pfam_id = $pfam_columns[0];
        $hash7{$columns[0]} = $pfam_columns[0];
        $hash7{$pfam_columns[0]} = $columns[0];
        $hash8{$pfam_columns[0]} = "$columns[0]\t$columns[12]\t$columns[11]";
    }
}

# read Pfam-A.clans.tsv
open my $pfamid2names, "/srv/db/pfam/27/Pfam-A.clans.tsv", or die "Couldn't open Pfam-A.clans.tsv\n";
open my $pfam_temp_OUT, ">pfam_output_temp.txt";
while (<$pfamid2names>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        if (exists $hash8{$columns[0]})
        {
                print {$pfam_temp_OUT} "$hash8{$columns[0]}\t$columns[4]\n";
        }
}

close($pfamoutput);
close($pfamid2names);
close($pfam_temp_OUT);

# hashes for tigrfam
my %hash9 = ();
my %hash10 = ();

# read tigrfam hmmscan output and store in hash
open my $tigrfamoutput, "./$locus.faaVStigr_all.hmm.hmmscanned.tab", or die "Couldn't open file $locus.faaVStigr_all.hmm.hmmscanned.tab\n";
while (<$tigrfamoutput>)
{
	next if /^\s*(#.*)?$/;
    next if $tigrfamoutput =~ /^#/;
    chomp $_;
    my @columns = split (/\t/, $_);
    if ($columns[5] > 10)
    {
        $hash9{$columns[2]} = $columns[0];
        $hash9{$columns[0]} = $columns[2];
        $hash10{$columns[0]} = "$columns[2]\t$columns[4]\t$columns[5]";
    }
}

# read tigrfam id2names2description
open my $tigrfamid2names, "/srv/db/tigrfam/14.0/TIGRFAMs_14.0_INFO/tigr_info_combined.parsed", or die "Couldn't open tigr_info_combined.parsed\n";
open my $tigrfam_temp_OUT, ">tigrfam_output_temp.txt";
while (<$tigrfamid2names>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
		$columns[0] =~ s/^\s+//;
		$columns[0] =~ s/\s+$//;
        if (exists $hash10{$columns[0]})
        {
                print {$tigrfam_temp_OUT} "$hash10{$columns[0]}\t$columns[1]\t$columns[2]\n";
        }
}

close($tigrfamoutput);
close($tigrfamid2names);
close($tigrfam_temp_OUT);

# hashes for cog
my %hash11 = ();
my %hash12 = ();
my %hash13 = ();

# read cog blastp output and store in hash
open my $cogblast, "./$locus.faaVSCOG.blastp", or die "Couldn't open file $locus.faaVSCOG.blastp\n";
while (<$cogblast>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        if ($columns[11] > 60)
        {
                $hash11{$columns[0]} = $columns[1];
                $hash11{$columns[1]} = $columns[0];
                $hash12{$columns[1]} = "$columns[0]\t$columns[10]\t$columns[11]";
        }
}

# read cog prot2COG.tab
open my $cogid2names, "/srv/db/cog/prot2COG.tab", or die "Couldn't open prot2COG.tab\n";
open my $cog_temp_OUT, "> cog_output_temp.txt";
while (<$cogid2names>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        if (exists $hash12{$columns[0]})
        {
                $hash13{$columns[0]} = "$hash12{$columns[0]}\t$columns[1]";
		$hash13{$columns[1]} = $hash12{$columns[0]};
                print {$cog_temp_OUT} "$hash12{$columns[0]}\t$columns[1]\n";
        }
}

close($cogblast);
close($cogid2names);
close($cog_temp_OUT);

# read cog listcogs.txt
open my $cogid2longernames, "/srv/db/cog/listcogs.txt", or die "Couldn't open listcogs.txt\n";
open my $cog_temp_OUT2, "> cog_output_temp2.txt";
while(<$cogid2longernames>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        if (exists $hash13{$columns[5]})
        {
                print {$cog_temp_OUT2} "$hash13{$columns[5]}\t$columns[3]\t$columns[4]\t$columns[6]\n";
        }
}

close($cog_temp_OUT2);


### now to parse all the temporary files and combine into one tab-delimited-file
# to store the IDs => DB => values/annotations
my %combined_bighash =();

# open file for output
open my $FINAL_OUTPUT, "> ./final_output.txt";
# print header
print {$FINAL_OUTPUT} "ORF_ID\timg_evalue\timg_score\timg_gene\timg_organism\timg_reciprocal\tuniref_evalue\tuniref_score\tuniref_gene\tuniref_organism\tuniref_reciprocal\tprokka_gene\tcog_evalues\tcog_scores\tcog_classes\tcog_gene_acronyms\tcog_genes\tpfam_evalues\tpfam_scores\tpfam_genes\ttigrfam_evalues\ttigrfam_scores\ttigrfam_genes\ttigrfam_descriptions\n";

# img
open my $img_annotation, "./img_output_temp2.txt", or die "Couldn't open img_output_temp2.txt\n";
while (<$img_annotation>)
{
	chomp $_;
	my @columns = split (/\t/, $_);
	my @baba = @columns[1..$#columns];
	#print "@baba \n"; 
	#$combined_bighash{$columns[0]}->{'img'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'a-img'}}, join("\t", @baba);
}

# uniref
open my $uniref_annotation, "./uniref_output_temp2.txt", or die "Couldn't open uniref_output_temp2.txt\n";
while (<$uniref_annotation>)
{
	chomp $_;
        my @columns = split (/\t/, $_);
	my @baba = @columns[1..$#columns];
	#$combined_bighash{$columns[0]}->{'uniref'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'b-uniref'}}, join("\t", @baba);
}

# prokka
# need to parse faa file to give prokka id2names
checkAndRunCommand("grep",[[
						"'>'",
						"prokka_annotation/$locus.faa |",
						"sed",
						"'s/>//g' |",
						"sed",
						-e => "'s/ /\\t/'",
						"> prokka_temp_output.txt",
						]], DIE_ON_FAILURE);

open my $prokka_annotation, "./prokka_temp_output.txt",  or die "Couldn't open prokka_temp_output.txt\n";
while (<$prokka_annotation>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        my @baba = @columns[1..$#columns];
       	#$combined_bighash{$columns[0]}->{'prokka'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'c-prokka'}}, join("\t", @baba);
}

# cog
open my $cog_annotation, "./cog_output_temp2.txt", or die "Couldn't open cog_output_temp2.txt\n";
while (<$cog_annotation>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        my @baba = @columns[1..$#columns];
	#$combined_bighash{$columns[0]}->{'cog'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'d-cog'}}, join("\t", @baba);
}

# pfam
open my $pfam_annotation, "./pfam_output_temp.txt", or die "Couldn't open pfam_output_temp.txt\n";
while (<$pfam_annotation>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        my @baba = @columns[1..$#columns];
        #$combined_bighash{$columns[0]}->{'pfam'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'e-pfam'}}, join("\t", @baba); 
}
#print Dumper \%combined_bighash;

# tigrfam
open my $tigrfam_annotation, "./tigrfam_output_temp.txt", or die "Couldn't open tigrfam_output_temp.txt\n";
while (<$tigrfam_annotation>)
{
        chomp $_;
        my @columns = split (/\t/, $_);
        my @baba = @columns[1..$#columns];
        #$combined_bighash{$columns[0]}->{'tigrfam'} = join("\t", @baba);
	push @{$combined_bighash{$columns[0]}->{'f-tigrfam'}}, join("\t", @baba);
}

# to print finally.................
# assign key and value for number of expected columns for each annotation type, important for putting NA in missing annotation types
my %column_lengths = (
    'a-img' => 5,
    'b-uniref' => 5, 
    'c-prokka' => 1,
    'd-cog' => 5,
    'e-pfam' => 3, 
    'f-tigrfam' => 4,
);

# print the orfids first 
foreach my $ID (sort(keys %combined_bighash))
{
        print {$FINAL_OUTPUT} "$ID\t";
        foreach my $annotation_type (sort(keys %column_lengths))
		{
		        # if the annotation type does not exist, print NA in the columns depending on the %column_lengths hash values
			if (! exists $combined_bighash{$ID}->{$annotation_type})
		        {	
		                # cool way of printing a certain string multiple times based on the values in the hash
				print {$FINAL_OUTPUT} join("\t", ("NA",) x $column_lengths{$annotation_type}), "\t";
		        }
			# check the derefencing made with @{$combined_bighash{$columns[0]}->{'f-tigrfam'}} and so on..
			# the derefencing allows the hash be converted into an array so that we can read the hash for the different types of annotation types
			elsif (ref($combined_bighash{$ID}->{$annotation_type}) eq 'ARRAY')
			{
			# place to store the columns in the values of the hash annotation types
                	my @storage_array;
  			foreach my $line (@{$combined_bighash{$ID}->{$annotation_type}}) 
			{
                        	# each annotation types have different number of columns, so we need to split the columns first before
				# we can add in the extra values if lets say an orfid hits multiple pfam/cog/tigrfam values
				my @values = split("\t",$line);
				# cool and alternate way of doing columns[1] = values[1], and so on.., repetitively 
				# what it basically means as long as the value i less than the number of columns in each annotation type
				# add +1 to the string $i and do the push below
				for (my $i = 0; $i <= $#values; $i++) 
				{
					push @{$storage_array[$i]}, $values[$i];
				}
			}
			#print Dumper(\@storage_array);
                	# array to store the multiple hits in each column.. for example test0001 orfid hits multiple pfam values like pf0008 anf pf0010 etc..
			# so we would like to have the values combined together in the same column delimited by a comma
			my @print_info_array;
			for (my $i = 0; $i < $column_lengths{$annotation_type}; $i++) 
			{
                        	push @print_info_array, join(", ", @{$storage_array[$i]});
                        }
                        #print Dumper(\@print_info_array);
                        print {$FINAL_OUTPUT} join("\t", @print_info_array), "\t";
			} 
			else
			{
          		print {$FINAL_OUTPUT} "$combined_bighash{$ID}{$annotation_type}\t";
 		       	}	
		}
	print {$FINAL_OUTPUT} "\n";	
}

#close all files
close($img_annotation);
close($uniref_annotation);
close($prokka_annotation);
close($cog_annotation);
close($pfam_annotation);
close($tigrfam_annotation);
close($FINAL_OUTPUT);


######################################################################
# CUSTOM SUBS
######################################################################

# who needs custom subs...

######################################################################
# TEMPLATE SUBS

######################################################################
# PARAMETERS

sub checkParams {
    #-----
    # Do any and all options checking here...
    #
    my @standard_options = ( "help|h+", "in|i:s", "locustag|l:s", "kingdom|k:s", "threads|t:s", "evalue|e:s");
    my %options;

    # Add any other command line options, and the code to handle them
    # 
    GetOptions( \%options, @standard_options );

    # if no arguments supplied print the usage and exit
    #
    exec("pod2usage $0") if (0 == (keys (%options) ));

    # If the -help option is set, print the usage and exit
    #
    exec("pod2usage $0") if $options{'help'};
    
    # Compulsory items
    #if(!exists $options{''} ) { printParamError (""); }
    if(!exists $options{'in'} ) { printParamError ("You MUST supply a fasta file"); }

    return \%options;
}

sub printParamError
{
    #-----
    # What to do if there's something wrong with a parameter
    #  
    my ($error) = @_;  
    print "**ERROR: $0 : $error\n"; exec("pod2usage $0");
}

sub overrideDefault
{
    #-----
    # Set and override default values for parameters
    #
    my ($default_value, $option_name) = @_;
    if(exists $global_options->{$option_name}) 
    {
        return $global_options->{$option_name};
    }
    return $default_value;
}

#####################################################################
# FILE IO

sub openWrite
{
    #-----
    # Open a file for writing
    #
    my ($fn) = @_;
    open my $fh, ">", $fn or croak "**ERROR: could not open file: $fn for writing $!\n";
    return $fh;
}

sub openRead
{   
    #-----
    # Open a file for reading
    #
    my ($fn) = @_;
    open my $fh, "<", $fn or croak "**ERROR: could not open file: $fn for reading $!\n";
    return $fh;
}

######################################################################
# EXTERNAL COMMANDS
#
# checkAndRunCommand("ls", {
#                          -a => ""
#                          }, 
#                          WARN_ON_FAILURE);

sub checkFileExists {
    #-----
    # Does a file exists?
    #
    my ($file) = @_;
    unless(-e $file) {
        croak "**ERROR: $0 : Cannot find:\n$file\n";
    }
}

sub logExternalCommand
{
    #-----
    # Log a command line command to the command line!
    #
    if(1 == $global_log_commands) {
        print $_[0], "\n";
    }
}

sub isCommandInPath
{
    #-----
    # Is this command in the path?
    #
    my ($cmd, $failure_type) = @_;
    if (system("which $cmd |> /dev/null")) {
        handleCommandFailure($cmd, $failure_type);
    }
}

sub runExternalCommand
{
    #-----
    # Run a command line command on the command line!
    #
    my ($cmd) = @_;
    logExternalCommand($cmd);
    system($cmd);
}

sub checkAndRunCommand
{
    #-----
    # Run external commands more sanelier
    #
    my ($cmd, $params, $failure_type) = @_;
    
    isCommandInPath($cmd, $failure_type);
    
    # join the parameters to the command
    my $param_str = join " ", map {formatParams($_)} @{$params};

    my $cmd_str = $cmd . " " . $param_str;
    
    print "The command currently running:\t$cmd_str\n"; 
    logExternalCommand($cmd_str);

    # make sure that all went well
    if (system($cmd_str)) {
         handleCommandFailure($cmd_str, $failure_type)
    }
}

sub formatParams {

    #---------
    # Handles and formats the different ways of passing parameters to 
    # checkAndRunCommand
    #
    my $ref = shift;
    
    if (ref($ref) eq "ARRAY") {
        return join(" ", @{$ref});
    } elsif (ref($ref) eq "HASH") {
       return join(" ", map { $_ . " " . $ref->{$_}} keys %{$ref});
    }
    croak 'The elements of the $params argument in checkAndRunCommand can ' .
        'only contain references to arrays or hashes\n';
}


sub handleCommandFailure {
    #-----
    # What to do when all goes bad!
    #
    my ($cmd, $failure_type) = @_;
    if (defined($failure_type)) {
        if ($failure_type == DIE_ON_FAILURE) {
            croak "**ERROR: $0 : " . $! . "\n";
        } elsif ($failure_type == WARN_ON_FAILURE) {
            carp "**WARNING: $0 : " . $! . "\n";
        }
    }
}


######################################################################
# MISC

sub printAtStart {
print<<"EOF";
---------------------------------------------------------------- 
 $0
 annotateM - annotate my genome
 Due to the blast processes against multiple databases, this whole
 annotation pipeline will usually take awhile. Please be patient!
 
---------------------------------------------------------------- 
EOF
}

__DATA__

=head1 NAME

   annotateM

=head1 COPYRIGHT

   Copyright (C) Mohamed Fauzi Haroon
   Special appearance from Adam Skarshewski

   This program is free software: you can redistribute it and/or modify
   it under the terms of the GNU General Public License as published by
   the Free Software Foundation, either version 3 of the License, or
   (at your option) any later version.

   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.

   You should have received a copy of the GNU General Public License
   along with this program.  If not, see <http://www.gnu.org/licenses/>.

=head1 DESCRIPTION

   Want to annotate your genome? annotateM!

=head1 SYNOPSIS

    annotateM -i [fasta_file] -l [locus] -k [Bacteria|Archaea] -t [threads] -e [evalue]

      -i FASTA_FILE				Nucleotide fasta file 
      -l locustag				Name of locus tag
      -k kingdom (Bacteria/Archaea)		Kingdom of genome to be annotated
      -t threads				Number of threads
      -e evalue					Evalue for BLAST, recommended 1e-3	
      [-help -h]				Displays basic usage information
         
=cut