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Summary

Bio::Tools::Run::StandAloneBlastPlus - Compute with NCBI's blast+ suite *ALPHA*

Package variables

Included modules

Inherit

Bio::Root::Root

Synopsis

NOTE: This module is related to the
Bio::Tools::Run::StandAloneBlast system in name (and inspiration)
only. You must use this module directly.

 # existing blastdb:
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb'
 );
 
 # create blastdb from fasta file and attach
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => 'myseqs.fas',
   -create => 1
 );
 
 # create blastdb from BioPerl sequence collection objects
 $alnio = Bio::AlignIO->new( -file => 'alignment.msf' );
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => $alnio,
   -create => 1
 );

 @seqs = $alnio->next_aln->each_seq;
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => \@seqs,
   -create => 1
 );

 # create database with masks

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
  -db_name => 'my_masked_db',
  -db_data => 'myseqs.fas',
  -masker => 'dustmasker',
  -mask_data => 'maskseqs.fas',
  -create => 1
 );

 # create a mask datafile separately
 $mask_file = $fac->make_mask(
   -data => 'maskseqs.fas',
   -masker => 'dustmasker'
 );

 # query database for metadata
 $info_hash = $fac->db_info;
 $num_seq = $fac->db_num_sequences;
 @mask_metadata = @{ $fac->db_filter_algorithms };

 # perform blast methods
 $result = $fac->tblastn( -query => $seqio );
 # see Bio::Tools::Run::StandAloneBlastPlus::BlastMethods 
 # for many more details

Description

NOTE: This module requires BLAST+ v. 2.2.24+ and higher. Until the API
stabilizes for BLAST+, consider this module highly experimental.
This module along with
Bio::Tools::Run::StandAloneBlastPlus::BlastMethods allows the user
to perform BLAST functions using the external program suite blast+
(available at
ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST/), using
BioPerl objects and Bio::SearchIO facilities. This wrapper can
prepare BLAST databases as well as run BLAST searches. It can also be
used to run blast+ programs independently.
This module encapsulates object construction and production of
databases and masks. Blast analysis methods (blastp, psiblast,
etc>) are contained in
Bio::Tools::Run::StandAloneBlastPlus::BlastMethods.

Methods

Methods description

 Title   : new
 Usage   : my $obj = new Bio::Tools::Run::StandAloneBlastPlus();
 Function: Builds a new Bio::Tools::Run::StandAloneBlastPlus object
 Returns : an instance of Bio::Tools::Run::StandAloneBlastPlus
 Args    : named argument (key => value) pairs:
           -db : blastdb name

 Title   : db
 Usage   : $obj->db($newval)
 Function: contains the basename of the local blast database
 Example : 
 Returns : value of db (a scalar string)
 Args    : readonly

 Title   : factory
 Usage   : $obj->factory($newval)
 Function: attribute containing the Bio::Tools::Run::BlastPlus 
           factory
 Example : 
 Returns : value of factory (Bio::Tools::Run::BlastPlus object)
 Args    : readonly

 Title   : program_version
 Usage   : $version = $bedtools_fac->program_version()
 Function: Returns the program version (if available)
 Returns : string representing location and version of the program
 Note    : this works around the WrapperBase::version() method conflicting with
           the -version parameter for SABlast (good argument for not having
           getter/setters for these)

 Title   : package_version
 Usage   : $version = $bedtools_fac->package_version()
 Function: Returns the BLAST+ package version (if available)
 Returns : string representing BLAST+ package version (may differ from version())

 Title   : make_db
 Usage   : 
 Function: create the blast database (if necessary), 
           imposing masking if specified
 Returns : true on success
 Args    :

 Title   : make_mask
 Usage   : 
 Function: create masking data based on specified parameters
 Returns : mask data filename (scalar string)
 Args    :

 Title   : db_info
 Usage   : 
 Function: get info for database 
           (via blastdbcmd -info); add factory attributes
 Returns : hash of database attributes
 Args    : [optional] db name (scalar string) (default: currently attached db)

 Title   : set_db_make_args
 Usage   : 
 Function: set the DB make arguments attribute 
           with checking
 Returns : true on success
 Args    : arrayref or hashref of named arguments

 Title   : set_mask_make_args
 Usage   : 
 Function: set the masker make arguments attribute
           with checking
 Returns : true on success
 Args    : arrayref or hasref of named arguments

 Title   : check_db
 Usage   : 
 Function: determine if database with registered name and dir
           exists
 Returns : 1 if db present, 0 if not present, undef if name/dir not
           set
 Args    : [optional] db name (default is 'registered' name in $self->db)
           [optional] db directory (default is 'registered' dir in 
                                    $self->db_dir)

 Title   : no_throw_on_crash
 Usage   : $fac->no_throw_on_crash($newval)
 Function: set to prevent an exeception throw on a failed 
           blast program execution
 Example : 
 Returns : value of no_throw_on_crash (boolean)
 Args    : on set, new value (boolean)

 Title   : _fastize
 Usage   : 
 Function: convert a sequence collection to a temporary
           fasta file (sans gaps)
 Returns : fasta filename (scalar string)
 Args    : sequence collection

 Title   : _register_temp_for_cleanup
 Usage   : 
 Function: register a file for cleanup with 
           cleanup() method
 Returns : true on success
 Args    : a file name or a blastdb basename
           (scalar string)

 Title   : cleanup
 Usage   : 
 Function: unlink files registered for cleanup
 Returns : true on success
 Args    :

 $fac->stderr

 $fac->factory->stderr

 $fac->db_num_sequences

Methods code

sub new {

    my ($class,@args) = @_;
    my $self = $class->SUPER::new(@args);
    my ($db_name, $db_data, $db_dir, $db_make_args,
	$mask_file, $mask_data, $mask_make_args, $masker, 
	$create, $overwrite, $is_remote, $prog_dir, $program_dir) 
                 = $self->_rearrange([qw( 
                                          DB_NAME
                                          DB_DATA
                                          DB_DIR
                                          DB_MAKE_ARGS
                                          MASK_FILE 
                                          MASK_DATA
                                          MASK_MAKE_ARGS
                                          MASKER
                                          CREATE
                                          OVERWRITE
                                          REMOTE
                                          PROG_DIR
                                          PROGRAM_DIR
                                           )], @args);

    # parm taint checks
    if ($db_name) {
	$self->throw("DB name contains invalid characters") unless $db_name =~ m{^[a-z0-9_/:.+-]+$}i;
    }

    if ( $db_dir ) { 
	$self->throw("DB directory (DB_DIR) not found") unless (-d $db_dir);
	$self->{'_db_dir'} = $db_dir;
    }
    else {
	$self->{'_db_dir'} = '.';
    }
    $program_dir ||= $prog_dir; # alias
    # now handle these systematically (bug #3003)
    # allow db_name to include path info
    # let db_dir act as root if present and db_name is a relative path
    # db property contains the pathless name only
    if ($db_name) {
	my ($v,$d,$f) = File::Spec->splitpath($db_name);
	$self->throw("No DB name at the end of path '$db_name'") unless $f;
	$f =~ s/\..*$//; # tolerant of extensions, but ignore them
	$self->{_db} = $f;
	# now establish db_path property as the internal authority on 
	# db location...
	if ( File::Spec->file_name_is_absolute($db_name) ) {
	    $self->throw("Path specified in DB name ('$d') does not exist") unless !$d || (-d $d);
	    $self->{_db_path} = File::Spec->catfile($d,$f);
	    $self->{_db_dir} = $d;
	    # ignore $db_dir, give heads-up
	    $self->warn("DB name is an absolute path; setting db_dir to '".$self->db_dir."'") if $db_dir;
	}
	else {
	    $d = File::Spec->catdir($self->db_dir, $d);
	    $self->throw("Path specified by DB_DIR+DB_NAME ('$d') does not exist") unless !$d || (-d $d);
	    $self->{_db_path} = File::Spec->catfile($d,$f);
	}
    }
	
    if ($masker) {
	$self->throw("Masker '$masker' not available") unless 
	    grep /^$masker$/, keys %AVAILABLE_MASKERS;
	$self->{_masker} = $masker;
    }
    
    if ($program_dir) {
	$self->throw("Can't find program directory '$program_dir'") unless
	    -d $program_dir;
	$self->{_program_dir} = $program_dir;
    }
    elsif ($ENV{BLASTPLUSDIR}) {
	$self->{_program_dir} = $ENV{BLASTPLUSDIR};
    }
    $Bio::Tools::Run::BlastPlus::program_dir = $self->{_program_dir} || 
	$Bio::Tools::Run::BlastPlus::program_dir;	
	

    $self->set_db_make_args( $db_make_args) if ( $db_make_args );
    $self->set_mask_make_args( $mask_make_args) if ($mask_make_args);
    $self->{'_create'} = $create;
    $self->{'_overwrite'} = $overwrite;
    $self->{'_is_remote'} = $is_remote;
    $self->{'_db_data'} = $db_data;
    
    $self->{'_mask_file'} = $mask_file;
    $self->{'_mask_data'} = $mask_data;


    # check db
    if (defined $self->check_db and $self->check_db == 0 and !$self->is_remote) {
	$self->throw("DB '".$self->db."' can't be found. To create, set -create => 1.") unless ($create || $overwrite);
    }
    if (!$self->db) {
	# allow this to pass; catch lazily at make_db...
	if (!$self->db_data) {
	    $self->debug('No database or db data specified. '.
			 'To create a new database, provide '.
			 '-db_data => [fasta|\@seqs|$seqio_object]')
	}

	# no db specified; create temp db
	$self->{_create} = 1;
	if ($self->db_dir) {
	    my $fh = File::Temp->new(TEMPLATE => 'DBXXXXX',
				     DIR => $self->db_dir,
				     UNLINK => 1);
	    my ($v,$d,$f) = File::Spec->splitpath($fh->filename);
	    $self->{_db} = $f;
	    $self->{_db_path} = $fh->filename;
	    $self->_register_temp_for_cleanup($self->db_path);
	    $fh->close;
	}
	else {
	    $self->{_db_dir} = File::Temp->newdir('DBDXXXXX');
	    $self->{_db} = 'DBTEMP';
	    $self->{_db_path} = File::Spec->catfile($self->db_dir, 
						    $self->db);
	}
    }

    return $self;

}

sub db {

 shift->{_db}

}

sub db_name {

 shift->{_db}

}

sub set_db_name {

 shift->{_db} = shift

}

sub db_dir {

 shift->{_db_dir}

}

sub set_db_dir {

 shift->{_db_dir} = shift

}

sub db_path {

 shift->{_db_path}

}

sub db_data {

 shift->{_db_data}

}

sub set_db_data {

 shift->{_db_data} = shift

}

sub db_type {

 shift->{_db_type}

}

sub masker {

 shift->{_masker}

}

sub set_masker {

 shift->{_masker} = shift

}

sub mask_file {

 shift->{_mask_file}

}

sub set_mask_file {

 shift->{_mask_file} = shift

}

sub mask_data {

 shift->{_mask_data}

}

sub set_mask_data {

 shift->{_mask_data} = shift

}

sub factory {

 shift->{_factory}

}

sub create {

 shift->{_create}

}

sub overwrite {

 shift->{_overwrite}

}

sub is_remote {

 shift->{_is_remote}

}

sub program_version {

    my $self = shift;
    my $fac = $self->factory;
    $fac->program_version(@_) if $fac;

}

sub package_version {

    my $self = shift;
    my $fac = $self->factory;
    $fac->package_version(@_) if $fac;

}

sub make_db {

    my $self = shift;
    my @args = @_;
    return 1 if ( $self->check_db && !$self->overwrite ); # already there or force make

    $self->throw('No database or db data specified. '.
		 'To create a new database, provide '.
		 '-db_data => [fasta|\@seqs|$seqio_object]') 
	unless $self->db_data;
    # db_data can be: fasta file, array of seqs, Bio::SeqIO object
    my $data = $self->db_data;
    $data = $self->_fastize($data);
    my $testio = Bio::SeqIO->new(-file=>$data, -format=>'fasta');
    $self->{_db_type} = ($testio->next_seq->alphabet =~ /.na/) ? 'nucl' : 'prot';
    $testio->close;

    my ($v,$d,$name) = File::Spec->splitpath($data);
    $name =~ s/\.fas$//;
    $self->{_db} ||= $name;
    $self->{_db_path} = File::Spec->catfile($self->db_dir,$self->db);
    # <#######[
    # deal with creating masks here, 
    # and provide correct parameters to the 
    # makeblastdb ...
    
    # accomodate $self->db_make_args here -- allow them
    # to override defaults, or allow only those args
    # that are not specified here?
    my $usr_db_args ||= $self->db_make_args;
    my %usr_args = @$usr_db_args if $usr_db_args;

    my %db_args = (
	-in => $data,
	-dbtype => $self->db_type,
	-out => $self->db_path,
	-title => $self->db,
	-parse_seqids => 1 # necessary for masking
	);
    # usr arg override
    if (%usr_args) {
	$db_args{$_} = $usr_args{$_} for keys %usr_args;
    }

    # do masking if requested
    # if the (masker and mask_data) OR mask_file attributes of this
    # object are set, assume that masking is desired
    # 
    if ($self->mask_file) { # the actual masking data is provided
	$db_args{'-mask_data'} = $self->mask_file;
    }
    elsif ($self->masker && $self->mask_data) { # build the mask
	$db_args{'-mask_data'} = $self->make_mask(-data => $self->mask_data);
	$self->throw("Masker error: message is '".$self->stderr."'") unless
	    $db_args{'-mask_data'};
	$self->{_mask_data} = $db_args{'-mask_data'};
    }

    $self->{_factory} = $bp_class->new(
	-command => 'makeblastdb',
	%db_args
	);
    $self->factory->no_throw_on_crash($self->no_throw_on_crash);    
    return $self->factory->_run;

}

sub make_mask {

    my $self = shift;
    my @args = @_;
    my ($data, $mask_db, $make_args, $masker) = $self->_rearrange([qw(
                                                            DATA
                                                            MASK_DB
                                                            MAKE_ARGS
                                                            MASKER)], @args);
    my (%mask_args,%usr_args,$db_type);
    my $infmt = 'fasta';
    $self->throw("make_mask requires -data argument") unless $data;
    $masker ||= $self->masker;
    $self->throw("no masker specified and no masker default set in object") 
	unless $masker;
    my $usr_make_args ||= $self->mask_make_args;
    %usr_args = @$usr_make_args if $usr_make_args;
    unless (grep /^$masker$/, keys %AVAILABLE_MASKERS) {
	$self->throw("Masker '$masker' not available");
    }
    if ($self->check_db($data)) {
	unless ($masker eq 'segmasker') {
	    $self->throw("Masker '$masker' can't use a blastdb as primary input");
	}
	unless ($self->db_info($data)->{_db_type} eq 
		$AVAILABLE_MASKERS{$masker}) {
	    $self->throw("Masker '$masker' is incompatible with input db sequence type");
	}
	$infmt = 'blastdb';
    }
    else {
	$data = $self->_fastize($data);
	my $sio = Bio::SeqIO->new(-file=>$data);
	my $s = $sio->next_seq;
	my $type;
	if ($s->alphabet =~ /.na/) {
	    $type = 'nucl';
	}
	elsif ($s->alphabet =~ /protein/) {
	    $type = 'prot';
	}
	else {
	    $type = 'UNK';
	}
	unless ($type eq $AVAILABLE_MASKERS{$masker}) {
	    $self->throw("Masker '$masker' is incompatible with sequence type '$type'");
	}
    }
    
    # check that sequence type and masker program match:
    
    # now, need to provide reasonable default masker arg settings, 
    # and override these with $usr_make_args as necessary and appropriate
    my $mh = File::Temp->new(TEMPLATE=>'MSKXXXXX',
			     UNLINK => 0,
			     DIR => $self->db_dir);
    my $mask_outfile = $mh->filename;
    $mh->close;
    $self->_register_temp_for_cleanup(File::Spec->catfile($self->db_dir,$mask_outfile));

    %mask_args = (
	-in => $data,
	-parse_seqids => 1,
	#-outfmt => $MASKER_ENCODING{$masker}
	);
    # usr arg override
    if (%usr_args) {
	$mask_args{$_} = $usr_args{$_} for keys %usr_args;
    }
    # masker-specific pipelines
    my $status;
    for ($masker) {
	m/dustmasker/ && do {
	    $mask_args{'-out'} = $mask_outfile;
	    $self->{_factory} = $bp_class->new(-command => $masker,
					       %mask_args);
	    $self->factory->no_throw_on_crash($self->no_throw_on_crash);
	    $status = $self->factory->_run;
	    last;
	};
	m/windowmasker/ && do {
	    # check mask_db if present
	    if ($mask_db) {
		unless ($self->check_db($mask_db)) {
		    $self->throw("Mask database '$mask_db' is not present or valid");
		}
	    }
	    my $cth = File::Temp->new(TEMPLATE=>'MCTXXXXX',
				      DIR => $self->db_dir);
	    my $ct_file = $cth->filename;
	    $cth->close;
	    $mask_args{'-out'} = $ct_file;
	    $mask_args{'-mk_counts'} = 'true';
	    $self->{_factory} = $bp_class->new(-command => $masker,
					       %mask_args);
	    $self->factory->no_throw_on_crash($self->no_throw_on_crash);
	    $status = $self->factory->_run;
	    last unless $status;
	    delete $mask_args{'-mk_counts'};
	    $mask_args{'-ustat'} = $ct_file;
	    $mask_args{'-out'} = $mask_outfile;
	    if ($mask_db) {
		$mask_args{'-in'} = $mask_db;
		$mask_args{'-infmt'} = 'blastdb';
	    }
	    $self->factory->reset_parameters(%mask_args);
	    $self->factory->no_throw_on_crash($self->no_throw_on_crash);
	    $status = $self->factory->_run;
	    last;
	};
	m/segmasker/ && do {
	    $mask_args{'-infmt'} = $infmt;
	    $mask_args{'-out'} = $mask_outfile;
	    $self->{_factory} = $bp_class->new(-command => $masker,
					       %mask_args);
	    $self->factory->no_throw_on_crash($self->no_throw_on_crash);
	    $status = $self->factory->_run;
	    last;
	};
	do {
	    $self->throw("Masker program '$masker' not recognized");
	};
    }
    return $status ? $mask_outfile : $status;

}

sub db_info {

    my $self = shift;
    my $db = shift;
    $db ||= $self->db_path;
    unless ($db) {
	$self->warn("db_info: db not specified and no db attached");
	return;
    }
    if ($self->is_remote) {
	$self->warn("db_info: sorry, can't get info for remote database (complain to NCBI)");
	return;
    }
    if ($db eq $self->db and $self->{_db_info}) {
	return $self->{_db_info}; # memoized
    }
    my $db_info_text;
    $self->{_factory} = $bp_class->new( -command => 'blastdbcmd',
					-info => 1,
					-db => $db );
    $self->factory->no_throw_on_crash(1);
    $self->factory->_run();
    $self->factory->no_throw_on_crash(0);
    if ($self->factory->stderr =~ /No alias or index file found/) {
	$self->warn("db_info: Couldn't find database ".$self->db."; make with make_db()");
	return;
    }
    $db_info_text = $self->factory->stdout;
    # parse info into attributes
    my $infh = IO::String->new($db_info_text);
    my %attr;
    while (<$infh>) {
	/Database: (.*)/ && do {
	    $attr{db_info_name} = $1;
	    next;
	};
	/([0-9,]+) sequences; ([0-9,]+) total/ && do {
	    $attr{db_num_sequences} = $1;
	    $attr{db_total_bases} = $2;
	    $attr{db_num_sequences} =~ s/,//g;
	    $attr{db_total_bases} =~ s/,//g;
	    next;
	};
	/Date: (.*?)\s+Longest sequence: ([0-9,]+)/ && do {
	    $attr{db_date} = $1; # convert to more usable date object
	    $attr{db_longest_sequence} = $2;
	    $attr{db_longest_sequence} =~ s/,//g;
	    next;
	};
	/Algorithm ID/ && do {
	    my $alg = $attr{db_filter_algorithms} = [];
	    while (<$infh>) {
		if (/\s+([0-9]+)\s+([a-z0-9_]+)\s+(.*)/i) {
		    push @$alg, { algorithm_id => $1,
				  algorithm_name => $2,
				  algorithm_opts => $3 };
		}
		else {
		    last;
		}
	    }
	    next;
	};
    }
    # get db type
    if ( -e $db.'.psq' ) {
	$attr{_db_type} = 'prot';
    }
    elsif (-e $db.'.nsq') {
	$attr{_db_type} = 'nucl';
    }
    else {
	$attr{_db_type} = 'UNK'; # bork
    }
    if ($db eq $self->db) {
	$self->{_db_type} = $attr{_db_type};
	$self->{_db_info_text} = $db_info_text;
	$self->{_db_info} =\% attr;
    }
    return\% attr;

}

sub set_db_make_args {

    my $self = shift;
    my $args = shift;
    $self->throw("Arrayref or hashref required at DB_MAKE_ARGS") unless 
	ref($args) =~ /^ARRAY|HASH$/;
    if (ref($args) eq 'HASH') {
	my @a = %$args;
	$args =\@ a;
    }
    $self->throw("Named args required for DB_MAKE_ARGS") unless !(@$args % 2);
    $self->{'_db_make_args'} = $args;
    return 1;

}

sub set_mask_make_args {

    my $self = shift;
    my $args = shift;
    $self->throw("Arrayref or hashref required at MASK_MAKE_ARGS") unless 
	ref($args) =~ /^ARRAY|HASH$/;
    if (ref($args) eq 'HASH') {
	my @a = %$args;
	$args =\@ a;
    }
    $self->throw("Named args required at MASK_MAKE_ARGS") unless !(@$args % 2);
    $self->{'_mask_make_args'} = $args;
    return 1;

}

sub check_db {

    my $self = shift;
    my ($db) = @_;
    my $db_path;
    if ($db) {
	my ($v,$d,$f) = File::Spec->splitpath($db);
	$f =~ s/\..*$//; # ignore extensions
	$db_path = File::Spec->catfile($d||'.',$f);
    }
    else {
	$db_path = $self->db_path;
    }
    if ( $db_path ) {
	$self->{_factory} = $bp_class->new( -command => 'blastdbcmd',
					    -info => 1,
					    -db => $db_path );
#	$DB::single=1;
	$self->factory->no_throw_on_crash(1);
	$self->factory->_run();
	$self->factory->no_throw_on_crash(0);
	return 0 if ($self->factory->stderr =~ /No alias or index file found/);
	return 1;
    }
    return;

}

sub no_throw_on_crash {

    my $self = shift;
    
    return $self->{'no_throw_on_crash'} = shift if @_;
    return $self->{'no_throw_on_crash'};

}

sub _fastize {

    my $self = shift;
    my $data = shift;
    for ($data) {
	!ref && do {
	    # suppose a fasta file name
	    $self->throw('Sequence file not found') unless -e $data;
	    my $guesser = Bio::Tools::GuessSeqFormat->new(-file => $data);
	    $self->throw('Sequence file not in FASTA format') unless
		$guesser->guess eq 'fasta';
	    last;
	};
	(ref eq 'ARRAY') && (ref $$data[0]) &&
	    ($$data[0]->isa('Bio::Seq') || $$data[0]->isa('Bio::PrimarySeq'))
	    && do {
		my $fh = File::Temp->new(TEMPLATE => 'DBDXXXXX', 
					 UNLINK => 0, 
					 DIR => $self->db_dir,
					 SUFFIX => '.fas');
		my $fname = $fh->filename;
		$fh->close;
		$self->_register_temp_for_cleanup($fname);
		my $fasio = Bio::SeqIO->new(-file=>">$fname", -format=>"fasta")
		   or $self->throw("Can't create temp fasta file");
		for (@$data) {
		    my $s = $_->seq;
		    my $a = $_->alphabet;
		    $s =~ s/[$Bio::PrimarySeq::GAP_SYMBOLS]//g;
		    $_->seq( $s );
		    $_->alphabet($a);
		    $fasio->write_seq($_);
		}
		$fasio->close;
		$data = $fname;
		last;
	};
	ref && do { # some kind of object
	    my ($fmt) = ref($data) =~ /.*::(.*)/;
	    if ($fmt eq 'fasta') {
		$data = $data->file; # use the fasta file directly
	    }
	    else {
		# convert
		my $fh = File::Temp->new(TEMPLATE => 'DBDXXXXX', 
					 UNLINK => 0, 
					 DIR => $self->db_dir,
					 SUFFIX => '.fas');
		my $fname = $fh->filename;
		$fh->close;
		$self->_register_temp_for_cleanup($fname);
		my $fasio = Bio::SeqIO->new(-file=>">$fname", -format=>"fasta") 
		    or $self->throw("Can't create temp fasta file");
		require Bio::PrimarySeq;
		if ($data->isa('Bio::AlignIO')) {
		    my $aln = $data->next_aln;
		    for ($aln->each_seq) {
			# must de-gap
			my $s = $_->seq;
			my $a = $_->alphabet;
			$s =~ s/[$Bio::PrimarySeq::GAP_SYMBOLS]//g;
			$_->seq( $s );
			$_->alphabet($a);
			$fasio->write_seq($_) 
		    }
		}
		elsif ($data->isa('Bio::SeqIO')) {
		    while (local $_ = $data->next_seq) {
			my $s = $_->seq;
			my $a = $_->alphabet;
			$s =~ s/[$Bio::PrimarySeq::GAP_SYMBOLS]//g;
			$_->seq( $s );
			$_->alphabet($a);
			$fasio->write_seq($_);
		    }
		}
		elsif ($data->isa('Bio::Align::AlignI')) {
		    for( $data->each_seq) {
			my $s = $_->seq;
			my $a = $_->alphabet;
			$s =~ s/[$Bio::PrimarySeq::GAP_SYMBOLS]//g;
			$_->seq( $s );
			$_->alphabet($a);
			$fasio->write_seq($_) 
		    }
		}
		elsif ($data->isa('Bio::Seq') || $data->isa('Bio::PrimarySeq')) {
		    my $s = $data->seq;
		    my $a = $data->alphabet;
		    $s =~ s/[$Bio::PrimarySeq::GAP_SYMBOLS]//g;
		    $data->seq($s);
		    $data->alphabet($a);
		    $fasio->write_seq($data);
		}
		else {
		    $self->throw("Can't handle sequence container object ".
				 "of type '".ref($data)."'");
		}
		$fasio->close;
		$data = $fname;
	    }
	    last;
	};
    }
    return $data;

}

sub _register_temp_for_cleanup {

    my $self = shift;
    my @files = @_;

    for (@files) {
	my ($v, $d, $n) = File::Spec->splitpath($_);
	$_ = File::Spec->catfile($self->db_dir, $n) unless length($d);
	push @{$self->{_cleanup_list}}, File::Spec->rel2abs($_);
    }
    return 1;

}

sub cleanup {

    my $self = shift;
    return unless $self->{_cleanup_list};
    for (@{$self->{_cleanup_list}}) {
	m/(\.[a-z0-9_]+)+$/i && do {
	    unlink $_;
	    next;
	};
	do { # catch all index files
	    if ( -e $_.".psq" ) {
		unlink glob($_.".p*");
		unlink glob($_.".??.p*");
	    }
	    elsif ( -e $_.".nsq" ) {
		unlink glob($_.".n*");
		unlink glob($_.".??.n*");
	    }
	    else {
		unlink $_;
	    }
	    next;
	};
    }
    return 1;

}

sub AUTOLOAD {

    my $self = shift;
    my @args = @_;
    my $method = $AUTOLOAD;
    $method =~ s/.*:://;
    my @ret;
    if (grep /^$method$/, @Bio::Tools::Run::StandAloneBlastPlus::BlastMethods) {
	push @args, ('-method_args' => ['-remote' => 1] ) if ($self->is_remote);
	return $self->run( -method => $method, @args );
    }
    if ($self->factory and $self->factory->can($method)) { # factory method
	return $self->factory->$method(@args);
    }
    if ($self->db_info and grep /^$method$/, keys %{$self->db_info}) {
	return $self->db_info->{$method};
    }
    # else, fail
    $self->throw("Can't locate method '$method' in class ".ref($self));

}


1;

}

General documentation

The basic mantra is to (1) create a BlastPlus factory using the
new() constructor, and (2) perform BLAST analyses by calling the
desired BLAST program by name off the factory object. The blast
database itself and any masking data are attached to the factory
object (step 1). Query sequences and any parameters associated with
particular programs are provided to the blast method call (step 2),
and are run against the attached database.

The factory needs to be told where the blast+ programs live. The
BLASTPLUSDIR environment variable will be checked for the default
executable directory. The program directory can be set for individual
factory instances with the PROG_DIR parameter. All the blast+
programs must be accessible from that directory (i.e., as executable
files or symlinks).
Either the database or BLAST subject data must be specified at object
construction. Databases can be pre-existing formatted BLAST dbs, or
can be built directly from fasta sequence files or BioPerl sequence
object collections of several kinds. The key constructor parameters
are DB_NAME, DB_DATA, DB_DIR.
To specify a pre-existing BLAST database, use DB_NAME alone:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
     -DB_NAME => 'mydb'
 );

The directory can be specified along with the basename, or separately
with DB_DIR:

$fac = Bio::Tools::Run::StandAloneBlastPlus->new(
-DB_NAME => '~/home/blast/mydb'
);

 #same as

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
     -DB_NAME => 'mydb', -DB_DIR => '~/home/blast'
 );

To create a BLAST database de novo, see /Creating a BLAST database.
If you wish to apply pre-existing mask data (i.e., the final ASN1
output from one of the blast+ masker programs), to the database before
querying, specify it with MASK_FILE:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
     -DB_NAME => 'mydb', -MASK_FILE => 'mymaskdata.asn'
 );

There are several options for creating the database de novo using
attached data, both before and after factory construction. If a
temporary database (one that can be deleted by the cleanup()
method) is desired, leave out the -db_name parameter. If
-db_name is specified, the database will be preserved with the
basename specified.
Use -create = 1> to create a new database (otherwise the factory
will look for an existing database). Use -overwrite = 1> to create
and overwrite an existing database.
Note that the database is not created immediately on factory
construction. It will be created if necessary on the first use of a
factory BLAST method, or you can force database creation by executing

 $fac->make_db();

    With a FASTA file:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => 'myseqs.fas',
   -create => 1
 );

    With another BioPerl object collection:

 $alnio = Bio::AlignIO->new( -file => 'alignment.msf' );
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => $alnio,
   -create => 1
 );
 @seqs = $alnio->next_aln->each_seq;
 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'mydb',
   -db_data => \@seqs,
   -create => 1
 );

    Other collections (e.g., Bio::SeqIO) are valid. If a certain type
does not work, please submit an enhancement request.
    To create temporary databases, leave out the -db_name, e.g.

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_data => 'myseqs.fas',
   -create => 1
 );

    To get the tempfile basename, do:

 $dbname = $fac->db;

    Use the explicit attribute setters:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -create => 1
 );

 $fac->set_db_data('myseqs.fas');
 $fac->make_db;

The blast+ mask utilities windowmasker, segmasker, and
dustmasker are available. Masking can be rolled into database
creation, or can be executed later. If your mask data is already
created and in ASN1 format, set the -mask_file attribute on
construction (see /Factory constuction/initialization).
To create a mask from raw data or an existing database and apply the
mask upon database creation, construct the factory like so:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'my_masked_db',
   -db_data => 'myseqs.fas',
   -masker => 'dustmasker',
   -mask_data => 'maskseqs.fas',
   -create => 1
 );

The masked database will be created during make_db.
The -mask_data parameter can be a FASTA filename or any BioPerl
sequence object collection. If the datatype ('nucl' or 'prot') of the
mask data is not compatible with the selected masker, an exception
will be thrown with a message to that effect.
To create a mask ASN1 file that can be used in the -mask_file
parameter separately from the attached database, use the
make_mask() method directly:

 $mask_file = $fac->make_mask(-data => 'maskseqs.fas',
                              -masker => 'dustmasker');
 # segmasker can use a blastdb as input
 $mask_file = $fac->make_mask(-mask_db => 'mydb',
                              -masker => 'segmasker')

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'my_masked_db',
   -db_data => 'myseqs.fas',
   -mask_file => $mask_file
   -create => 1
 );

To get a hash containing useful metadata on an existing database
(obtained by running blastdbcmd -info, use db_info():

# get info on the attached database..
$info = $fac->db_info;
# get info on another database
$info = $fac->db_info('~/home/blastdbs/another');
To get a particular info element for the attached database, just call
the element name off the factory:

 $num_seqs = $fac->db_num_sequences;
 # info on all the masks applied to the db, if any:
 @masking_info = @{ $fac->db_filter_algorithms };

The blast+ programs are actually executed by a
Bio::Tools::Run::BlastPlus wrapper instance. This instance is
available for peeking and poking in the StandAloneBlastPlus
factory() attribute. For convenience, BlastPlus methods can be
run from the StandAloneBlastPlus object, and are delegated to the
factory() attribute. For example, to get the blast+ program to be
executed, examine either

 $fac->factory->command

or

 $fac->command

Similarly, the current parameters for the BlastPlus factory are

 @parameters = $fac->get_parameters

Temporary analysis files produced under a single factory instances can
be unlinked by running

 $fac->cleanup;

Tempfiles are generally not removed unless this method is explicitly
called. cleanup() only unlinks "registered" files and
databases. All temporary files are automatically registered; in
particular, "anonymous" databases (such as

 $fac->Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_data => 'myseqs.fas', 
   -create => 1
 );

without a -db_name specification) are registered for cleanup. Any
file or database can be registered with an internal method:

 $fac->_register_temp_for_cleanup('testdb');

    You can check whether a given basename points to a properly formatted
BLAST database by doing

 $is_good = $fac->check_db('putative_db');

    User parameters can be passed to the underlying blast+ programs (if
you know what you're doing) with db_make_args and mask_make_args:

 $fac = Bio::Tools::Run::StandAloneBlastPlus->new(
   -db_name => 'customdb',
   -db_data => 'myseqs.fas', 
   -db_make_args => [ '-taxid_map' => 'seq_to_taxa.txt' ],
   -masker => 'windowmasker',
   -mask_data => 'myseqs.fas',
   -mask_make_args => [ '-dust' => 'T' ],
   -create => 1
 );

    You can prevent exceptions from being thrown by failed blast+ program
executions by setting no_throw_on_crash. Examine the error with
stderr():

 $fac->no_throw_on_crash(1);
 $fac->make_db;
 if ($fac->stderr =~ /Error:/) {
    #handle error
    ...
 }

Bio::Tools::Run::StandAloneBlastPlus::BlastMethods,
Bio::Tools::Run::BlastPlus

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.

  bioperl-l@bioperl.org                  - General discussion
http://bioperl.org/wiki/Mailing_lists  - About the mailing lists

Please direct usage questions or support issues to the mailing list:
bioperl-l@bioperl.org
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.

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 web:

  http://redmine.open-bio.org/projects/bioperl/

Email maj -at- fortinbras -dot- us

The rest of the documentation details each of the object methods.
Internal methods are usually preceded with a _