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1 # $Id: ContigAnalysis.pm,v 1.2 2002/12/01 00:03:28 jason Exp $
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2 #
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3 # BioPerl module for Bio::Assembly::ContigAnalysis
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4 #
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5 # Cared for by Robson francisco de Souza <rfsouza@citri.iq.usp.br>
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6 #
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7 # Copyright Robson Francisco de Souza
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8 #
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9 # You may distribute this module under the same terms as perl itself
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10 #
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11 # POD documentation - main docs before the code
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12
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13 =head1 NAME
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14
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15 Bio::Assembly::ContigAnalysis -
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16 Perform analysis on sequence assembly contigs.
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17
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18 =head1 SYNOPSIS
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19
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20 # Module loading
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21 use Bio::Assembly::ContigAnalysis;
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22
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23 # Assembly loading methods
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24 my $ca = new Bio::Assembly::ContigAnalysis( -contig=>$contigOBJ );
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25
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26 my @lcq = $ca->low_consensus_quality;
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27 my @hqd = $ca->high_quality_discrepancies;
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28 my @ss = $ca->single_strand_regions;
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29
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30 =head1 DESCRIPTION
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31
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32 A contig is as a set of sequences, locally aligned to each other, when
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33 the sequences in a pair may be aligned. It may also include a
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34 consensus sequence. Bio::Assembly::ContigAnalysis is a module
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35 holding a collection of methods to analyze contig objects. It was
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36 developed around the Bio::Assembly::Contig implementation of contigs and
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37 can not work with another contig interface.
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38
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39 =head1 FEEDBACK
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40
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41 =head2 Mailing Lists
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42
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43 User feedback is an integral part of the evolution of this and other
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44 Bioperl modules. Send your comments and suggestions preferably to the
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45 Bioperl mailing lists Your participation is much appreciated.
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46
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47 bioperl-l@bioperl.org - General discussion
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48 http://bio.perl.org/MailList.html - About the mailing lists
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49
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50 =head2 Reporting Bugs
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51
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52 Report bugs to the Bioperl bug tracking system to help us keep track
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53 the bugs and their resolution. Bug reports can be submitted via email
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54 or the web:
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55
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56 bioperl-bugs@bio.perl.org
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57 http://bugzilla.bioperl.org/
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58
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59 =head1 AUTHOR - Robson Francisco de Souza
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60
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61 Email: rfsouza@citri.iq.usp.br
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62
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63 =head1 APPENDIX
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64
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65 The rest of the documentation details each of the object
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66 methods. Internal methods are usually preceded with a _
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67
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68 =cut
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69
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70 package Bio::Assembly::ContigAnalysis;
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71
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72 use Bio::Root::Root;
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73 use strict;
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74 use vars qw(@ISA);
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75
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76 @ISA = qw(Bio::Root::Root);
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77
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78 =head1 Object creator
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79
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80 =head2 new
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81
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82 Title : new
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83 Usage : my $contig = Bio::Assembly::ContigAnalysis->new(-contig=>$contigOBJ);
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84 Function : Creates a new contig analysis object
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85 Returns : Bio::Assembly::ContigAnalysis
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86 Args :
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87 -contig : a Bio::Assembly::Contig object
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88
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89 =cut
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90
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91 sub new {
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92 my($class,@args) = @_;
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93 my $self = $class->SUPER::new(@args);
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94
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95 my ($contigOBJ) = $self->_rearrange([qw(CONTIG)],@args);
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96 unless ($contigOBJ->isa("Bio::Assembly::Contig")) {
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97 $self->throw("ContigAnal works only on Bio::Assembly::Contig objects\n");
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98 }
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99
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100 $self->{'_objref'} = $contigOBJ;
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101 return $self;
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102 }
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103
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104 =head1 Analysis methods
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105
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106 =head2 high_quality_discrepancies
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107
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108 Title : high_quality_discrepancies
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109 Usage : my $sfc = $ContigAnal->high_quality_discrepancies();
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110 Function :
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111
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112 Locates all high quality discrepancies among aligned
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113 sequences and the consensus sequence.
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114
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115 Note: see Bio::Assembly::Contig POD documentation,
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116 section "Coordinate System", for a definition of
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117 available types. Default coordinate system type is
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118 "gapped consensus", i.e. consensus sequence (with gaps)
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119 coordinates. If limits are not specified, the entire
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120 alignment is analyzed.
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121
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122 Returns : Bio::SeqFeature::Collection
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123 Args : optional arguments are
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124 -threshold : cutoff value for low quality (minimum high quality)
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125 Default: 40
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126 -ignore : number of bases that will not be analysed at
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127 both ends of contig aligned elements
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128 Default: 5
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129 -start : start of interval that will be analyzed
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130 -end : start of interval that will be analyzed
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131 -type : coordinate system type for interval
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132
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133 =cut
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134
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135 sub high_quality_discrepancies {
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136 my ($self,@args) = shift; # Package reference
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137
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138 my ($threshold,$ignore,$start,$end,$type) =
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139 $self->_rearrange([qw(THRESHOLD IGNORE START END TYPE)],@args);
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140
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141 # Defining default threhold and HQD_ignore
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142 $threshold = 40 unless (defined($threshold));
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143 $ignore = 5 unless (defined($ignore));
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144 $type = 'gapped consensus' unless (defined($type));
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145
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146 # Changing input coordinates system (if needed)
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147 if (defined $start && $type ne 'gapped consensus') {
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148 $start = $self->{'_objref'}->change_coord($type,'gapped consensus',$start);
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149 } elsif (!defined $start) {
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150 $start = 1;
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151 }
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152 if (defined $end && $type ne 'gapped consensus') {
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153 $end = $self->{'_objref'}->change_coord($type,'gapped consensus',$end);
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154 } elsif (!defined $end) {
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155 $end = $self->{'_objref'}->get_consensus_length();
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156 }
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157
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158 # Scanning each read sequence and the contig sequence and
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159 # adding discrepancies to Bio::SeqFeature::Collection
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160 my @seqIDs = $self->{'_objref'}->get_seq_ids(-start=>$start,
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161 -end=>$end,
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162 -type=>$type);
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163 my $consensus = $self->{'_objref'}->get_consensus_sequence()->seq;
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164
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165 my @HQD = ();
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166 foreach my $seqID (@seqIDs) {
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167 # Setting aligned read sub-sequence limits and loading data
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168 my $seq = $self->{'_objref'}->get_seq_by_name($seqID);
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169 my $qual = $self->{'_objref'}->get_qual_by_name($seqID);
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170 unless (defined $qual) {
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171 $self->warn("Can't correctly evaluate HQD without aligned sequence qualities for $seqID");
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172 next;
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173 }
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174 my $sequence = $seq->seq;
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175 my @quality = @{ $qual->qual };
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176
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177 # Scanning the aligned region of each read
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178 my $seq_ix = 0;
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179 my $coord = $self->{'_objref'}->get_seq_feat_by_tag($seq,"_align_clipping:$seqID");
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180 my ($astart,$aend) = ($coord->start,$coord->end);
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181 $astart = $astart + $ignore; # Redefining limits to evaluate HQDs (jump $ignore at start)
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182 $aend = $aend - $ignore; # Redefining limits to evaluate HQDs (stop $ignore before end)
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183
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184 my ($d_start,$d_end,$i);
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185 for ($i=$astart-1; $i<=$aend-1; $i++) {
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186 # Changing coordinate $i+1 from 'gapped consensus' mode to "aligned $seqID" (coordinate $seq_ix)
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187 $seq_ix = $self->{'_objref'}->change_coord('gapped consensus',"aligned $seqID",$i+1);
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188 next unless (($i >= $start) && ($i <= $end));
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189
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190 my $r_base = uc(substr($sequence,$seq_ix-1,1));
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191 my $c_base = uc(substr($consensus,$i,1));
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192
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193 # Discrepant region start: store $d_start and $type
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194 (!defined($d_start) &&
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195 ($r_base ne $c_base) &&
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196 ($quality[$seq_ix-1] >= $threshold)) && do {
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197 $d_start = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i+1);
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198 #print $seqID," ",$r_base," ",$i+1," ",$c_base," ",$contig_ix-1," ",$quality[$i]," $type\n";
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199 next;
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200 };
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201
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202 # Quality change or end of discrepant region: store limits and undef $d_start
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203 if (defined($d_start) &&
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204 (($quality[$seq_ix-1] < $threshold) ||
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205 (uc($r_base) eq uc($c_base)))) {
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206 $d_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i);
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207 #print $seqID," ",$r_base," ",$i+1," ",$c_base," ",$contig_ix-1," ",$quality[$i]," $type\n";
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208 push(@HQD, Bio::SeqFeature::Generic->new(-primary=>"high_quality_discrepancy:$seqID",
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209 -start=>$d_start,
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210 -end=>$d_end,
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211 -strand=>$seq->strand()) );
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212 $d_start = undef;
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213 next;
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214 }
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215 } # for ($i=$astart-1; $i<=$aend-1; $i++)
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216
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217 # Loading discrepancies located at sub-sequence end, if any.
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218 if (defined($d_start)) {
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219 $d_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i);
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220 push(@HQD, Bio::SeqFeature::Generic->new(-primary=>"high_quality_discrepancy:$seqID",
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221 -start=>$d_start,
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222 -end=>$d_end,
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223 -strand=>$seq->strand()) );
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224 }
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225 } # foreach my $seqID (@seqIDs)
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226
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227 return @HQD;
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228 }
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229
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230 =head2 low_consensus_quality
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231
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232 Title : low_consensus_quality
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233 Usage : my $sfc = $ContigAnal->low_consensus_quality();
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234 Function : Locates all low quality regions in the consensus
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235 Returns : an array of Bio::SeqFeature::Generic objects
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236 Args : optional arguments are
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237 -threshold : cutoff value for low quality (minimum high quality)
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238 Default: 25
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239 -start : start of interval that will be analyzed
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240 -end : start of interval that will be analyzed
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241 -type : coordinate system type for interval
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242
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243 =cut
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244
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245 sub low_consensus_quality {
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246 my ($self,@args) = shift; # Packege reference
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247
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248 my ($threshold,$start,$end,$type) =
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249 $self->_rearrange([qw(THRESHOLD START END TYPE)],@args);
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250
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251 # Setting default value for threshold
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252 $threshold = 25 unless (defined($threshold));
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253
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254 # Loading qualities
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255 my @quality = @{ $self->{'_objref'}->get_consensus_quality()->qual };
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256
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257 # Changing coordinates to gap mode noaln (consed: consensus without alignments)
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258 $start = 1 unless (defined($start));
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259 if (defined $start && defined $type && ($type ne 'gapped consensus')) {
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260 $start = $self->{'objref'}->change_coord($type,'gapped consensus',$start);
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261 $end = $self->{'objref'}->change_coord($type,'gapped consensus',$end) if (defined($end));
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262 }
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263 $end = $self->{'_objref'}->get_consensus_length unless (defined $end);
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264
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265 # Scanning @quality vector and storing intervals limits with base qualities less then
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266 # the threshold value
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267 my ($lcq_start);
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268 my ($i,@LCQ);
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269 for ($i=$start-1; $i<=$end-1; $i++) {
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270 # print $quality[$i],"\t",$i,"\n";
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271 if (!defined($lcq_start) && (($quality[$i] <= $threshold) || ($quality[$i] == 98))) {
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272 $lcq_start = $i+1;
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273 } elsif (defined($lcq_start) && ($quality[$i] > $threshold)) {
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274 $lcq_start = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$lcq_start);
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275 my $lcq_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i);
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276 push(@LCQ, Bio::SeqFeature::Generic->new(-start=>$lcq_start,
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277 -end=>$lcq_end,
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278 -primary=>'low_consensus_quality') );
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279 $lcq_start = undef;
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280 }
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281 }
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282
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283 if (defined $lcq_start) {
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284 $lcq_start = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$lcq_start);
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285 my $lcq_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i);
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286 push(@LCQ, Bio::SeqFeature::Generic->new(-start=>$lcq_start,
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287 -end=>$lcq_end,
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288 -primary=>'low_consensus_quality') );
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289 }
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290
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291 return @LCQ;
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292 }
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293
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294 =head2 not_confirmed_on_both_strands
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295
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296 Title : low_quality_consensus
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297 Usage : my $sfc = $ContigAnal->low_quality_consensus();
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298 Function :
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299
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300 Locates all regions whose consensus bases were not
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301 confirmed by bases from sequences aligned in both
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302 orientations, i.e., in such regions, no bases in aligned
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303 sequences of either +1 or -1 strand agree with the
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304 consensus bases.
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305
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306 Returns : an array of Bio::SeqFeature::Generic objects
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307 Args : optional arguments are
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308 -start : start of interval that will be analyzed
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309 -end : start of interval that will be analyzed
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310 -type : coordinate system type for interval
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311
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312 =cut
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313
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314 sub not_confirmed_on_both_strands {
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315 my ($self,@args) = shift; # Package reference
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316
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317 my ($start,$end,$type) =
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318 $self->_rearrange([qw(START END TYPE)],@args);
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319
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320 # Changing coordinates to default system 'align' (contig sequence with alignments)
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321 $start = 1 unless (defined($start));
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322 if (defined($type) && ($type ne 'gapped consensus')) {
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323 $start = $self->{'_objref'}->change_coord($type,'gapped consensus',$start);
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324 $end = $self->{'_objref'}->change_coord($type,'gapped consensus',$end) if (defined($end));
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325 }
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326 $end = $self->{'_objref'}->get_consensus_length unless (defined($end));
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327
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328 # Scanning alignment
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329 my %confirmed = (); # If ($confirmed{$orientation}[$i] > 0) then $i is confirmed in $orientation strand
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330 my ($i);
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331 my $consensus = $self->{'_objref'}->get_consensus_sequence()->seq;
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332 foreach my $seqID ($self->{'_objref'}->get_seq_ids) {
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333 # Setting aligned read sub-sequence limits and loading data
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334 my $seq = $self->{'_objref'}->get_seq_by_name($seqID);
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335 my $sequence = $seq->seq;
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336
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337 # Scanning the aligned regions of each read and registering confirmed sites
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338 my $contig_ix = 0;
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339 my $coord = $self->{'_objref'}->get_seq_feat_by_tag($seq,"_align_clipping:$seqID");
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340 my ($astart,$aend,$orientation) = ($coord->start,$coord->end,$coord->strand);
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341 $astart = $self->{'_objref'}->change_coord('gapped consensus',"aligned $seqID",$astart);
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342 $aend = $self->{'_objref'}->change_coord('gapped consensus',"aligned $seqID",$aend);
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343 for ($i=$astart-1; $i<=$aend-1; $i++) {
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344 # $i+1 in 'align' mode is $contig_ix
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345 $contig_ix = $self->{'_objref'}->change_coord("aligned $seqID",'gapped consensus',$i+1);
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346 next unless (($contig_ix >= $start) && ($contig_ix <= $end));
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347 my $r_base = uc(substr($sequence,$i,1));
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348 my $c_base = uc(substr($consensus,$contig_ix-1,1));
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349 if ($c_base eq '-') {
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350 $confirmed{$orientation}[$contig_ix] = -1;
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351 } elsif (uc($r_base) eq uc($c_base)) { # Non discrepant region found
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352 $confirmed{$orientation}[$contig_ix]++;
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353 }
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354 } # for ($i=$astart-1; $i<=$aend-1; $i++)
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355 } # foreach $seqID (@reads)
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356
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357 # Locating non-confirmed aligned regions for each orientation in $confirmed registry
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358 my ($orientation);
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359 my @NCBS = ();
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360 foreach $orientation (keys %confirmed) {
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361 my ($ncbs_start,$ncbs_end);
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362
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363 for ($i=$start; $i<=$end; $i++) {
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364 if (!defined($ncbs_start) &&
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365 (!defined($confirmed{$orientation}[$i]) || ($confirmed{$orientation}[$i] == 0))) {
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366 $ncbs_start = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i);
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367 } elsif (defined($ncbs_start) &&
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368 defined($confirmed{$orientation}[$i]) &&
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369 ($confirmed{$orientation}[$i] > 0)) {
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370 $ncbs_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$i-1);
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371 push(@NCBS, Bio::SeqFeature::Generic->new(-start=>$ncbs_start,
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372 -end=>$ncbs_end,
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373 -strand=>$orientation,
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374 -primary=>"not_confirmed_on_both_strands") );
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375 $ncbs_start = undef;
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376 }
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377 }
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378
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379 if (defined($ncbs_start)) { # NCBS at the end of contig
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380 $ncbs_end = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$end);
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381 push(@NCBS, Bio::SeqFeature::Generic->new(-start=>$ncbs_start,
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382 -end=>$ncbs_end,
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383 -strand=>$orientation,
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384 -primary=>'not_confirmed_on_both_strands') );
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385 }
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386 }
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387
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388 return @NCBS;
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389 }
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390
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391 =head2 single_strand
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392
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393 Title : single_strand
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394 Usage : my $sfc = $ContigAnal->single_strand();
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395 Function :
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396
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397 Locates all regions covered by aligned sequences only in
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398 one of the two strands, i.e., regions for which aligned
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399 sequence's strand() method returns +1 or -1 for all
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400 sequences.
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401
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402 Returns : an array of Bio::SeqFeature::Generic objects
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403 Args : optional arguments are
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404 -start : start of interval that will be analyzed
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405 -end : start of interval that will be analyzed
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406 -type : coordinate system type for interval
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407
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408 =cut
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409
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410 #'
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411 sub single_strand {
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412 my ($self,@args) = shift; # Package reference
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413
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414 my ($start,$end,$type) =
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415 $self->_rearrange([qw(START END TYPE)],@args);
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416
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417 # Changing coordinates to gap mode align (consed: consensus sequence with alignments)
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418 $type = 'gapped consensus' unless(defined($type));
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419 $start = 1 unless (defined($start));
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420 if (defined($type) && $type ne 'gapped consensus') {
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421 $start = $self->{'objref'}->change_coord($type,'gapped consensus',$start);
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422 $end = $self->{'objref'}->change_coord($type,'gapped consensus',$end) if (defined($end));
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423 }
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424 ($end) = $self->{'_objref'}->get_consensus_length unless (defined($end));
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425
|
|
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426 # Loading complete list of coordinates for aligned sequences
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|
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427 my $sfc = $self->{'_objref'}->get_features_collection();
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428 my @forward = grep { $_->primary_tag =~ /^_aligned_coord:/ }
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|
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429 $sfc->features_in_range(-start=>$start,
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|
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430 -end=>$end,
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|
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431 -contain=>0,
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|
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432 -strand=>1,
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|
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433 -strandmatch=>'strong');
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|
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434 my @reverse = grep { $_->primary_tag =~ /^_aligned_coord:/ }
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|
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435 $sfc->features_in_range(-start=>$start,
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|
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436 -end=>$end,
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|
|
437 -contain=>0,
|
|
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438 -strand=>-1,
|
|
|
439 -strandmatch=>'strong');
|
|
|
440 # Merging overlapping features
|
|
|
441 @forward = $self->_merge_overlapping_features(@forward);
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|
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442 @reverse = $self->_merge_overlapping_features(@reverse);
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|
|
443
|
|
|
444 # Finding single stranded regions
|
|
|
445 my ($length) = $self->{'_objref'}->get_consensus_length;
|
|
|
446 $length = $self->{'_objref'}->change_coord('gapped consensus','ungapped consensus',$length);
|
|
|
447 @forward = $self->_complementary_features_list(1,$length,@forward);
|
|
|
448 @reverse = $self->_complementary_features_list(1,$length,@reverse);
|
|
|
449
|
|
|
450 my @SS = ();
|
|
|
451 foreach my $feat (@forward, @reverse) {
|
|
|
452 $feat->primary_tag('single_strand_region');
|
|
|
453 push(@SS,$feat);
|
|
|
454 }
|
|
|
455
|
|
|
456 return @SS;
|
|
|
457 }
|
|
|
458
|
|
|
459 =head1 Internal Methods
|
|
|
460
|
|
|
461 =head2 _merge_overlapping_features
|
|
|
462
|
|
|
463 Title : _merge_overlapping_features
|
|
|
464 Usage : my @feat = $ContigAnal->_merge_overlapping_features(@features);
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|
|
465 Function : Merge all overlapping features into features
|
|
|
466 that hold original features as sub-features
|
|
|
467 Returns : array of Bio::SeqFeature::Generic objects
|
|
|
468 Args : array of Bio::SeqFeature::Generic objects
|
|
|
469
|
|
|
470 =cut
|
|
|
471
|
|
|
472 sub _merge_overlapping_features {
|
|
|
473 my ($self,@feat) = @_;
|
|
|
474
|
|
|
475 $self->throw_not_implemented();
|
|
|
476 }
|
|
|
477
|
|
|
478 =head2 _complementary_features_list
|
|
|
479
|
|
|
480 Title : _complementary_features_list
|
|
|
481 Usage : @feat = $ContigAnal->_complementary_features_list($start,$end,@features);
|
|
|
482 Function : Build a list of features for regions
|
|
|
483 not covered by features in @features array
|
|
|
484 Returns : array of Bio::SeqFeature::Generic objects
|
|
|
485 Args :
|
|
|
486 $start : [integer] start of first output feature
|
|
|
487 $end : [integer] end of last output feature
|
|
|
488 @features : array of Bio::SeqFeature::Generic objects
|
|
|
489
|
|
|
490 =cut
|
|
|
491
|
|
|
492 sub _complementary_features_list {
|
|
|
493 my ($self,$start,$end,@feat) = @_;
|
|
|
494
|
|
|
495 $self->throw_not_implemented();
|
|
|
496 }
|
|
|
497
|
|
|
498 1;
|
|
|
499
|
|
|
500 __END__
|
