Bio::Assembly::Contig - Perl module to hold and manipulate sequence assembly contigs. # Module loading use Bio::Assembly::IO; # Assembly loading methods $aio = new Bio::Assembly::IO(-file=>"test.ace.1", -format=>'phrap'); $assembly = $aio->next_assembly; foreach $contig ($assembly->all_contigs) { # do something } # OR, if you want to build the contig yourself, use Bio::Assembly::Contig; $c = Bio::Assembly::Contig->new(-id=>"1"); $ls = Bio::LocatableSeq->new(-seq=>"ACCG-T", -id=>"r1", -alphabet=>'dna'); $ls2 = Bio::LocatableSeq->new(-seq=>"ACA-CG-T", -id=>"r2", -alphabet=>'dna'); $ls_coord = Bio::SeqFeature::Generic->new(-start=>3, -end=>8, -strand=>1); $ls2_coord = Bio::SeqFeature::Generic->new(-start=>1, -end=>8, -strand=>1); $c->add_seq($ls); $c->add_seq($ls2); $c->set_seq_coord($ls_coord,$ls); $c->set_seq_coord($ls2_coord,$ls2); $con = Bio::LocatableSeq->new(-seq=>"ACACCG-T", -alphabet=>'dna'); $c->set_consensus_sequence($con); $l = $c->change_coord('unaligned r2','ungapped consensus',6); print "6 in unaligned r2 => $l in ungapped consensus\n"; A contig is as a set of sequences, locally aligned to each other, so that every sequence has overlapping regions with at least one sequence in the contig, such that a continuous of overlapping sequences is formed, allowing the deduction of a consensus sequence which may be longer than any of the sequences from which it was deduced. In this documentation we refer to the overlapping sequences used to build the contig as "aligned sequences" and to the sequence deduced from the overlap of aligned sequences as the "consensus". Methods to deduce the consensus sequence from aligned sequences were not yet implemented in this module, but its posssible to add a consensus sequence deduced by other means, e.g, by the assembly program used to build the alignment. All aligned sequences in a Bio::Assembly::Contig must be Bio::Assembly::Locatable objects and have a unique ID. The unique ID restriction is due to the nature of the module's internal data structures and is also a request of some assembly programs. If two sequences with the same ID are added to a contig, the first sequence added is replaced by the second one.

Coordinate_systems
There are four base coordinate systems in Bio::Assembly::Contig. When you need to access contig elements or data that exists on a certain range or location, you may be specifying coordinates in relation to different sequences, which may be either the contig consensus or one of the aligned sequences that were used to do the assembly. ========================================================= Name | Referenced sequence --------------------------------------------------------- "gapped consensus" | Contig (with gaps) "ungapped consensus" | Contig (without gaps) "aligned $seqID" | sequence $seqID (with gaps) "unaligned $seqID" | sequence $seqID (without gaps) ========================================================= "gapped consensus" refers to positions in the aligned consensus sequence, which is the consensus sequence including the gaps inserted to align it agains the aligned sequences that were used to assemble the contig. So, its limits are [ 1, (consensus length + number of gaps in consensus) ] "ungapped consensus" is a coordinate system based on the consensus sequence, but excluding consensus gaps. This is just the coordinate system that you have when considering the consensus sequence alone, instead of aligned to other sequences. "aligned $seqID" refers to locations in the sequence $seqID after alignment of $seqID against the consensus sequence (reverse complementing the original sequence, if needed). Coordinate 1 in "aligned $seqID" is equivalent to the start location (first base) of $seqID in the consensus sequence, just like if the aligned sequence $seqID was a feature of the consensus sequence. "unaligned $seqID" is equivalent to a location in the isolated sequence, just like you would have when considering the sequence alone, out of an alignment. When changing coordinates from "aligned $seq2" to "unaligned $seq2", if $seq2 was reverse complemented when included in the alignment, the output coordinates will be reversed to fit that fact, i.e. 1 will be changed to length($seq2), 2 will be length($seq)-1 and so on. An important note: when you change gap coordinates from a gapped system ("gapped consensus" or "aligned $seqID") to a system that does not include gaps ("ungapped consensus" or "unaligned $seqID"), the position returned will be the first location before all gaps neighboring the input location.

Feature_collection
Bio::Assembly::Contig stores much information about a contig in a Bio::Assembly::SeqFeature::Collection object. Relevant information on the alignment is accessed by selecting features based on their primary tags (e.g. all features which have a primary tag of the form '_aligned_coord:$seqID', where $seqID is an aligned sequence ID, are coordinates for sequences in the contig alignment) and, by using methods from Bio::Assembly::SeqFeature::Collection, it's possible to select features by overlap with other features. We suggest that you use the primary tags of features as identifiers for feature classes. By convention, features with primary tags starting with a '_' are generated by modules that populate the contig data structure and return the contig object, maybe as part of an assembly object, e.g. drivers from the Bio::Assembly::IO set. Features in the features collection may be associated with particular aligned sequences. To obtain this, you must attach the sequence to the feature, using attach() seq from Bio::Assembly::SeqFeatureI, before you add the feature to the feature collection. We also suggest to add the sequence id to the primary tag, so that is easy to select feature for a particular sequence. There is only one feature class that some methods in Bio::Assembly::Contig expect to find in the feature collection: features with primary tags of the form '_aligned_coord:$seqID', where $seqID is the aligned sequence id (like returned by $seq->id()). These features describe the position (in "gapped consensus" coordinates) of aligned sequences, and the method set_seq_coord() automatically changes a feature's primary tag to this form whenever the feature is added to the collection by this method. Only two methods in Bio::Assembly::Contig will not work unless there are features from this class: change_coord() and get_seq_coord(). Other feature classes will be automatically available only when Bio::Assembly::Contig objects are created by a specific module. Such feature classes are (or should be) documented in the documentation of the module which create them, to which the user should refer.

Mailing Lists
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 lists Your participation is much appreciated. bioperl-l@bioperl.org - General discussion http://bio.perl.org/MailList.html - About the mailing lists

Reporting Bugs
Report bugs to the Bioperl bug tracking system to help us keep track the bugs and their resolution. Bug reports can be submitted via email or the web: bioperl-bugs@bio.perl.org http://bugzilla.bioperl.org/ rfsouza@citri.iq.usp.br The rest of the documentation details each of the object methods. Internal methods are usually preceded with a _

new
Title : new Usage : my $contig = new Bio::Assembly::Contig(); Function : Creates a new contig object Returns : Bio::Assembly::Contig Args : -source => string representing the source program where this contig came from -id => contig unique ID These methods exist to enable adding information about possible relations among contigs, e.g. when you already have a scaffold for your assembly, describing the ordering of contigs in the final assembly, but no sequences covering the gaps between neighboring contigs.

source
Title : source Usage : $contig->source($program); Function : Get/Set program used to build this contig Returns : string Argument : [optional] string

assembly
Title : assembly Usage : $contig->assembly($assembly); Function : Get/Set assembly object for this contig Returns : a Bio::Assembly::Scaffold object Argument : a Bio::Assembly::Scaffold object

strand
Title : strand Usage : $contig->strand($num); Function : Get/Set contig orientation in a scaffold/assembly. Its equivalent to the strand property of sequence objects and sets whether the contig consensus should be reversed and complemented before being added to a scaffold or assembly. Returns : integer Argument : 1 if orientaion is forward, -1 if reverse and 0 if none

upstream_neighbor
Title : upstream_neighbor Usage : $contig->upstream_neighbor($contig); Function : Get/Set a contig neighbor for the current contig when building a scaffold. The upstream neighbor is located before $contig first base Returns : nothing Argument : Bio::Assembly::Contig

downstream_neighbor
Title : downstream_neighbor Usage : $contig->downstream_neighbor($num); Function : Get/Set a contig neighbor for the current contig when building a scaffold. The downstream neighbor is located after $contig last base Returns : nothing Argument : Bio::Assembly::Contig

add_features
Title : add_features Usage : $contig->add_features($feat,$flag) Function : Add an array of features to the contig feature collection. The consensus sequence may be attached to the added feature, if $flag is set to 1. If $flag is 0 and the feature attached to one of the contig aligned sequences, the feature is registered as an aligned sequence feature. If $flag is 0 and the feature is not attched to any sequence in the contig, the feature is simply added to the feature collection and no attachment or registration is made. Note: You must attach aligned sequences to their features prior to calling add_features, otherwise you won't be able to access the feature through get_seq_feat_by_tag() method. Returns : number of features added. Argument : $feat : A reference to an array of Bio::SeqFeatureI $flag : boolean - true if consensus sequence object should be attached to this feature, false if no consensus attachment should be made. Default: false.

remove_features
Title : remove_features Usage : $contig->remove_features(@feat) Function : Remove an array of contig features Returns : number of features removed. Argument : An array of Bio::SeqFeatureI

get_features_collection
Title : get_features_collection Usage : $contig->get_features_collection() Function : Get the collection of all contig features Returns : Bio::SeqFeature::Collection Argument : none See Coordinate_Systems above.

change_coord
Title : change_coord Usage : $contig->change_coord($in,$out,$query) Function : Change coordinate system for $query. This method transforms locations between coordinate systems described in section "Coordinate Systems" of this document. Note: this method will throw an exception when changing coordinates between "ungapped consensus" and other systems if consensus sequence was not set. It will also throw exceptions when changing coordinates among aligned sequence, either with or without gaps, and other systems if sequence locations were not set with set_seq_coord(). Returns : integer Argument : $in : [string] input coordinate system $out : [string] output coordinate system $query : [integer] a position in a sequence

get_seq_coord
Title : get_seq_coord Usage : $contig->get_seq_coord($seq); Function : Get "gapped consensus" location for aligned sequence Returns : Bio::SeqFeature::Generic for coordinates or undef. A warning is printed if sequence coordinates were not set. Argument : Bio::LocatabaleSeq object

set_seq_coord
Title : set_seq_coord Usage : $contig->set_seq_coord($feat,$seq); Function : Set "gapped consensus" location for an aligned sequence. If the sequence was previously added using add_seq, its coordinates are changed/set. Otherwise, add_seq is called and the sequence is added to the contig. Returns : Bio::SeqFeature::Generic for old coordinates or undef. Argument : $feat : a Bio::SeqFeature::Generic object representing a location for the aligned sequence, in "gapped consensus" coordinates. Note: the original feature primary tag will be lost. $seq : a Bio::LocatabaleSeq object

set_consensus_sequence
Title : set_consensus_sequence Usage : $contig->set_consensus_sequence($seq) Function : Set the consensus sequence object for this contig Returns : consensus length Argument : Bio::LocatableSeq

set_consensus_quality
Title : set_consensus_quality Usage : $contig->set_consensus_quality($qual) Function : Set the quality object for consensus sequence Returns : nothing Argument : Bio::Seq::QualI object

get_consensus_length
Title : get_consensus_length Usage : $contig->get_consensus_length() Function : Get consensus sequence length Returns : integer Argument : none

get_consensus_sequence
Title : get_consensus_sequence Usage : $contig->get_consensus_sequence() Function : Get a reference to the consensus sequence object for this contig Returns : Bio::SeqI object Argument : none

get_consensus_quality
Title : get_consensus_quality Usage : $contig->get_consensus_quality() Function : Get a reference to the consensus quality object for this contig. Returns : A Bio::QualI object Argument : none

set_seq_qual
Title : set_seq_qual Usage : $contig->set_seq_qual($seq,$qual); Function : Adds quality to an aligned sequence. Returns : nothing Argument : a Bio::LocatableSeq object and a Bio::Seq::QualI object See Bio::LocatableSeq for more information.

get_seq_ids
Title : get_seq_ids Usage : $contig->get_seq_ids(-start=>$start, -end=>$end, -type=>"gapped A0QR67B08.b"); Function : Get list of sequence IDs overlapping inteval [$start, $end] The default interval is [1,$contig->length] Default coordinate system is "gapped contig" Returns : An array Argument : A hash with optional elements: -start : consensus subsequence start -end : consensus subsequence end -type : the coordinate system type for $start and $end arguments Coordinate system avaliable are: "gapped consensus" : consensus coordinates with gaps "ungapped consensus" : consensus coordinates without gaps "aligned $ReadID" : read $ReadID coordinates with gaps "unaligned $ReadID" : read $ReadID coordinates without gaps

get_seq_feat_by_tag
Title : get_seq_feat_by_tag Usage : $seq = $contig->get_seq_feat_by_tag($seq,"_aligned_coord:$seqID") Function : Get a sequence feature based on its primary_tag. When you add Returns : a Bio::SeqFeature object Argument : a Bio::LocatableSeq and a string (feature primary tag)

get_seq_by_name
Title : get_seq_by_name Usage : $seq = $contig->get_seq_by_name('Seq1') Function : Gets a sequence based on its id. Returns : a Bio::LocatableSeq object undef if name is not found Argument : string

get_qual_by_name
Title : get_qual_by_name Usage : $seq = $contig->get_qual_by_name('Seq1') Function : Gets Bio::Seq::QualI object for a sequence through its id ( as given by $qual->id() ). Returns : a Bio::Seq::QualI object. undef if name is not found Argument : string

Modifier methods
These methods modify the MSE by adding, removing or shuffling complete sequences.

add_seq
Title : add_seq Usage : $contig->add_seq($newseq); Function : Adds a sequence to the contig. *Does* *not* align it - just adds it to the hashes. Returns : nothing Argument : a Bio::LocatableSeq object See Bio::LocatableSeq for more information.

remove_seq
Title : remove_seq Usage : $contig->remove_seq($seq); Function : Removes a single sequence from an alignment Returns : 1 on success, 0 otherwise Argument : a Bio::LocatableSeq object

purge
Title : purge Usage : $contig->purge(0.7); Function: Removes sequences above whatever %id. This function will grind on large alignments. Beware! (perhaps not ideally implemented) Example : Returns : An array of the removed sequences Argument:

sort_alphabetically
Title : sort_alphabetically Usage : $contig->sort_alphabetically Function : Changes the order of the alignemnt to alphabetical on name followed by numerical by number. Returns : Argument :

Sequence selection methods
Methods returning one or more sequences objects.

each_seq
Title : each_seq Usage : foreach $seq ( $contig->each_seq() ) Function : Gets an array of Seq objects from the alignment Returns : an array Argument :

each_alphabetically
Title : each_alphabetically Usage : foreach $seq ( $contig->each_alphabetically() ) Function : Returns an array of sequence object sorted alphabetically by name and then by start point. Does not change the order of the alignment Returns : Argument :

each_seq_with_id
Title : each_seq_with_id Usage : foreach $seq ( $contig->each_seq_with_id() ) Function : Gets an array of Seq objects from the alignment, the contents being those sequences with the given name (there may be more than one) Returns : an array Argument : a seq name

get_seq_by_pos
Title : get_seq_by_pos Usage : $seq = $contig->get_seq_by_pos(3) Function : Gets a sequence based on its position in the alignment. Numbering starts from 1. Sequence positions larger than no_sequences() will thow an error. Returns : a Bio::LocatableSeq object Argument : positive integer for the sequence osition

Create new alignments
The result of these methods are horizontal or vertical subsets of the current MSE.

select
Title : select Usage : $contig2 = $contig->select(1, 3) # three first sequences Function : Creates a new alignment from a continuous subset of sequences. Numbering starts from 1. Sequence positions larger than no_sequences() will thow an error. Returns : a Bio::Assembly::Contig object Argument : positive integer for the first sequence positive integer for the last sequence to include (optional)

select_noncont
Title : select_noncont Usage : $contig2 = $contig->select_noncont(1, 3) # first and 3rd sequences Function : Creates a new alignment from a subset of sequences. Numbering starts from 1. Sequence positions larger than no_sequences() will thow an error. Returns : a Bio::Assembly::Contig object Args : array of integers for the sequences

slice
Title : slice Usage : $contig2 = $contig->slice(20, 30) Function : Creates a slice from the alignment inclusive of start and end columns. Sequences with no residues in the slice are excluded from the new alignment and a warning is printed. Slice beyond the length of the sequence does not do padding. Returns : a Bio::Assembly::Contig object Argument : positive integer for start column positive integer for end column

Change sequences within the MSE
These methods affect characters in all sequences without changeing the alignment.

map_chars
Title : map_chars Usage : $contig->map_chars('\.','-') Function : Does a s/$arg1/$arg2/ on the sequences. Useful for gap characters Notice that the from (arg1) is interpretted as a regex, so be careful about quoting meta characters (eg $contig->map_chars('.','-') wont do what you want) Returns : Argument : 'from' rexexp 'to' string

uppercase
Title : uppercase() Usage : $contig->uppercase() Function : Sets all the sequences to uppercase Returns : Argument :

match_line
Title : match_line() Usage : $contig->match_line() Function : Generates a match line - much like consensus string except that a line indicating the '*' for a match. Argument : (optional) Match line characters ('*' by default) (optional) Strong match char (':' by default) (optional) Weak match char ('.' by default)

match
Title : match() Usage : $contig->match() Function : Goes through all columns and changes residues that are identical to residue in first sequence to match '.' character. Sets match_char. USE WITH CARE: Most MSE formats do not support match characters in sequences, so this is mostly for output only. NEXUS format (Bio::AlignIO::nexus) can handle it. Returns : 1 Argument : a match character, optional, defaults to '.'

unmatch
Title : unmatch() Usage : $contig->unmatch() Function : Undoes the effect of method match. Unsets match_char. Returns : 1 Argument : a match character, optional, defaults to '.'

MSE attibutes
Methods for setting and reading the MSE attributes. Note that the methods defining character semantics depend on the user to set them sensibly. They are needed only by certain input/output methods. Unset them by setting to an empty string ('').

id
Title : id Usage : $contig->id("Ig") Function : Gets/sets the id field of the alignment Returns : An id string Argument : An id string (optional)

missing_char
Title : missing_char Usage : $contig->missing_char("?") Function : Gets/sets the missing_char attribute of the alignment It is generally recommended to set it to 'n' or 'N' for nucleotides and to 'X' for protein. Returns : An missing_char string, Argument : An missing_char string (optional)

match_char
Title : match_char Usage : $contig->match_char('.') Function : Gets/sets the match_char attribute of the alignment Returns : An match_char string, Argument : An match_char string (optional)

gap_char
Title : gap_char Usage : $contig->gap_char('-') Function : Gets/sets the gap_char attribute of the alignment Returns : An gap_char string, defaults to '-' Argument : An gap_char string (optional)

symbol_chars
Title : symbol_chars Usage : my @symbolchars = $contig->symbol_chars; Function: Returns all the seen symbols (other than gaps) Returns : array of characters that are the seen symbols Argument: boolean to include the gap/missing/match characters

Alignment descriptors
These read only methods describe the MSE in various ways.

consensus_string
Title : consensus_string Usage : $str = $contig->consensus_string($threshold_percent) Function : Makes a strict consensus Returns : Argument : Optional treshold ranging from 0 to 100. The consensus residue has to appear at least threshold % of the sequences at a given location, otherwise a '?' character will be placed at that location. (Default value = 0%)

consensus_iupac
Title : consensus_iupac Usage : $str = $contig->consensus_iupac() Function : Makes a consensus using IUPAC ambiguity codes from DNA and RNA. The output is in upper case except when gaps in a column force output to be in lower case. Note that if your alignment sequences contain a lot of IUPAC ambiquity codes you often have to manually set alphabet. Bio::PrimarySeq::_guess_type thinks they indicate a protein sequence. Returns : consensus string Argument : none Throws : on protein sequences

is_flush
Title : is_flush Usage : if( $contig->is_flush() ) : : Function : Tells you whether the alignment : is flush, ie all of the same length : : Returns : 1 or 0 Argument :

length
Title : length() Usage : $len = $contig->length() Function : Returns the maximum length of the alignment. To be sure the alignment is a block, use is_flush Returns : Argument :

maxdisplayname_length
Title : maxdisplayname_length Usage : $contig->maxdisplayname_length() Function : Gets the maximum length of the displayname in the alignment. Used in writing out various MSE formats. Returns : integer Argument :

no_residues
Title : no_residues Usage : $no = $contig->no_residues Function : number of residues in total in the alignment Returns : integer Argument :

no_sequences
Title : no_sequences Usage : $depth = $contig->no_sequences Function : number of sequence in the sequence alignment Returns : integer Argument : None

percentage_identity
Title : percentage_identity Usage : $id = $contig->percentage_identity Function: The function calculates the percentage identity of the alignment Returns : The percentage identity of the alignment (as defined by the implementation) Argument: None

overall_percentage_identity
Title : percentage_identity Usage : $id = $contig->percentage_identity Function: The function calculates the percentage identity of the conserved columns Returns : The percentage identity of the conserved columns Args : None

average_percentage_identity
Title : average_percentage_identity Usage : $id = $contig->average_percentage_identity Function: The function uses a fast method to calculate the average percentage identity of the alignment Returns : The average percentage identity of the alignment Args : None

Alignment positions
Methods to map a sequence position into an alignment column and back. column_from_residue_number() does the former. The latter is really a property of the sequence object and can done using Bio::LocatableSeq::location_from_column: # select somehow a sequence from the alignment, e.g. my $seq = $contig->get_seq_by_pos(1); #$loc is undef or Bio::LocationI object my $loc = $seq->location_from_column(5);

column_from_residue_number
Title : column_from_residue_number Usage : $col = $contig->column_from_residue_number( $seqname, $resnumber) Function: This function gives the position in the alignment (i.e. column number) of the given residue number in the sequence with the given name. For example, for the alignment Seq1/91-97 AC..DEF.GH Seq2/24-30 ACGG.RTY.. Seq3/43-51 AC.DDEFGHI column_from_residue_number( "Seq1", 94 ) returns 5. column_from_residue_number( "Seq2", 25 ) returns 2. column_from_residue_number( "Seq3", 50 ) returns 9. An exception is thrown if the residue number would lie outside the length of the aligment (e.g. column_from_residue_number( "Seq2", 22 ) Note: If the the parent sequence is represented by more than one alignment sequence and the residue number is present in them, this method finds only the first one. Returns : A column number for the position in the alignment of the given residue in the given sequence (1 = first column) Args : A sequence id/name (not a name/start-end) A residue number in the whole sequence (not just that segment of it in the alignment)

Sequence names
Methods to manipulate the display name. The default name based on the sequence id and subsequence positions can be overridden in various ways.

displayname
Title : displayname Usage : $contig->displayname("Ig", "IgA") Function : Gets/sets the display name of a sequence in the alignment : Returns : A display name string Argument : name of the sequence displayname of the sequence (optional)

set_displayname_count
Title : set_displayname_count Usage : $contig->set_displayname_count Function : Sets the names to be name_# where # is the number of times this name has been used. Returns : None Argument : None

set_displayname_flat
Title : set_displayname_flat Usage : $contig->set_displayname_flat() Function : Makes all the sequences be displayed as just their name, not name/start-end Returns : 1 Argument : None

set_displayname_normal
Title : set_displayname_normal Usage : $contig->set_displayname_normal() Function : Makes all the sequences be displayed as name/start-end Returns : None Argument : None

_binary_search
Title : _binary_search Usage : _binary_search($list,$query) Function : Find a number in a sorted list of numbers. Return values may be on or two integers. One positive integer or zero (>=0) is the index of the element that stores the queried value. Two positive integers (or zero and another number) are the indexes of elements among which the queried value should be placed. Negative single values mean: -1: $query is smaller than smallest element in list -2: $query is greater than greatest element in list Returns : array of integers Argument : $list : array reference $query : integer

_compare
Title : _compare Usage : _compare($arg1,$arg2) Function: Perform numeric or string comparisons Returns : integer (0, 1 or -1) Args : values to be compared

_nof_gaps
Title : _nof_gaps Usage : _nof_gaps($array_ref, $query) Function: number of gaps found before position $query Returns : integer Args : $array_ref : gap registry reference $query : [integer] a position in a sequence

_padded_unpadded
Title : _padded_unpadded Usage : _padded_unpadded($array_ref, $query) Function: Returns a coordinate corresponding to position $query after gaps were removed from a sequence. Returns : integer Args : $array_ref : reference to this gap registry $query : [integer] coordionate to change

_unpadded_padded
Title : _unpadded_padded Usage : _unpadded_padded($array_ref, $query) Function: Returns the value corresponding to ungapped position $query when gaps are counted as valid sites in a sequence Returns : Args : $array_ref = a reference to this sequence's gap registry $query = [integer] location to change

_register_gaps
Title : _register_gaps Usage : $self->_register_gaps($seq, $array_ref) Function: stores gap locations for a sequence Returns : number of gaps found Args : $seq : sequence string $array_ref : a reference to an array, where gap locations will be stored