Bio::Assembly::IO::sam - An IO module for assemblies in Sam format *BETA*
$aio = Bio::Assembly::IO( -file => "mysam.bam", -refdb => "myrefseqs.fas"); $assy = $aio->next_assembly;
This is a (currently) read-only IO module designed to convert Sequence/Alignment Map (SAM; http://samtools.sourceforge.net/) formatted alignments to Bio::Assembly::Scaffold representations, containing .Bio::Assembly::Contig and Bio::Assembly::Singlet objects. It uses lstein's Bio::DB::Sam to parse binary formatted SAM (.bam) files guided by a reference sequence fasta database.
NB: Bio::DB::Sam is not a BioPerl module; it can be obtained via CPAN. It in turn requires the libbam library; source can be downloaded at http://samtools.sourceforge.net/.
Bio::DB::Sam
libbam
Required files
A binary SAM (.bam) alignment and a reference sequence database in FASTA format are required. Various required indexes (.fai, .bai) will be created as necessary (via Bio::DB::Sam).
.bam
.fai
.bai
Compressed files
...can be specified directly , if IO::Uncompress::Gunzip is installed. Get it from your local CPAN mirror.
BAM vs. SAM
The input alignment should be in (possibly gzipped) binary SAM (.bam) format. If it isn't, you will get a message explaining how to convert it, viz.:
$ samtools view -Sb mysam.sam > mysam.bam
The bam file must also be sorted on coordinates: do
$ samtools sort mysam.unsorted.bam > mysam.bam
Contigs
Contigs are calculated by this module, using the 'coverage' feature of the Bio::DB::Sam object. A contig represents a contiguous portion of a reference sequence having non-zero coverage at each base.
The bwa assembler (http://bio-bwa.sourceforge.net/) can assign read sequences to multiple reference sequence locations. The present implementation currently assigns such reads only to the first contig in which they appear.
Consensus sequences
Consensus sequence and quality objects are calculated by this module, using the pileup callback feature of Bio::DB::Sam. The consensus is (currently) simply the residue at a position that has the maximum sum of quality values. The consensus quality is the integer portion of the simple average of quality values for the consensus residue.
pileup
SeqFeatures
Read sequences stored in contigs are accompanied by the following features:
contig : name of associated contig cigar : CIGAR string for this read
If the read is paired with a successfully mapped mate, these features will also be available:
mate_start : coordinate of to which the mate was aligned mate_len : length of mate read mate_strand : strand of mate (-1 or 1) insert_size : size of insert spanned by the mate pair
These features are obtained as follows:
@ids = $contig->get_seq_ids; $an_id = $id[0]; # or whatever $seq = $contig->get_seq_by_name($an_id); # Bio::LocatableSeq's aren't SeqFeature containers, so... $feat = $contig->get_seq_feat_by_tag( $seq, "_aligned_coord:".$s->id ); ($cigar) = $feat->get_tag_values('cigar'); # etc.
Supporting both text SAM (TAM) and binary SAM (BAM)
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:
https://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 _
Title : next_assembly Usage : my $scaffold = $asmio->next_assembly(); Function: return the next assembly in the sam-formatted stream Returns : Bio::Assembly::Scaffold object Args : none
Title : next_contig Usage : my $contig = $asmio->next_contig(); Function: return the next contig or singlet from the sam stream Returns : Bio::Assembly::Contig or Bio::Assembly::Singlet Args : none
Title : write_assembly Usage : Function: not implemented (module currrently read-only) Returns : Args :
Title : _store_contig Usage : my $contigobj = $self->_store_contig(\%contiginfo); Function: create and load a contig object Returns : Bio::Assembly::Contig object Args : Bio::DB::Sam::Segment object
Title : _store_read Usage : my $readobj = $self->_store_read($readobj, $contigobj); Function: store information of a read belonging to a contig in a contig object Returns : Bio::LocatableSeq Args : Bio::DB::Bam::AlignWrapper, Bio::Assembly::Contig
Title : _store_singlet Usage : my $singletobj = $self->_store_singlet($contigobj); Function: convert a contig object containing a single read into a singlet object Returns : Bio::Assembly::Singlet Args : Bio::Assembly::Contig (previously loaded with only one seq)
Title : _init_sam Usage : $self->_init_sam($fasfile) Function: obtain a Bio::DB::Sam parsing of the associated sam file Returns : true on success Args : [optional] name of the fasta reference db (scalar string) Note : The associated file can be plain text (.sam) or binary (.bam); If the fasta file is not specified, and no file is contained in the refdb() attribute, a .fas file with the same basename as the sam file will be searched for.
Title : _get_contig_segs_from_coverage Usage : Function: calculates separate contigs using coverage info in the segment Returns : array of Bio::DB::Sam::Segment objects, representing each contig Args : Bio::DB::Sam::Segment object
Title : _calc_consensus_quality Usage : @qual = $aio->_calc_consensus_quality( $contig_seg ); Function: calculate an average or other data-reduced quality over all sites represented by the features contained in a Bio::DB::Sam::Segment Returns : Args : a Bio::DB::Sam::Segment object
Title : _calc_consensus Usage : @qual = $aio->_calc_consensus( $contig_seg ); Function: calculate a simple quality-weighted consensus sequence for the segment Returns : a SeqWithQuality object Args : a Bio::DB::Sam::Segment object
Title : refdb Usage : $obj->refdb($newval) Function: the name of the reference db fasta file Example : Returns : value of refdb (a scalar) Args : on set, new value (a scalar or undef, optional)
Title : _segset Usage : $segset_hashref = $self->_segset() Function: hash container for the Bio::DB::Sam::Segment objects that represent each set of contigs for each seq_id { $seq_id => [@contig_segments], ... } Example : Returns : value of _segset (a hashref) if no arg, or the arrayref of contig segments, if arg == a seq id Args : [optional] seq id (scalar string) Note : readonly; hash elt set in _init_sam()
Title : _current_refseq_id Usage : $obj->_current_refseq_id($newval) Function: the "current" reference sequence id Example : Returns : value of _current_refseq (a scalar) Args : on set, new value (a scalar or undef, optional)
Title : current_contig_seg_idx Usage : $obj->current_contig_seg_idx($newval) Function: the "current" segment index in the "current" refseq Example : Returns : value of current_contig_seg_idx (a scalar) Args : on set, new value (a scalar or undef, optional)
Title : sam Usage : $obj->sam($newval) Function: store the associated Bio::DB::Sam object Example : Returns : value of sam (a Bio::DB::Sam object) Args : on set, new value (a scalar or undef, optional)
To install Bio::Seq, copy and paste the appropriate command in to your terminal.
cpanm
cpanm Bio::Seq
CPAN shell
perl -MCPAN -e shell install Bio::Seq
For more information on module installation, please visit the detailed CPAN module installation guide.