NAME

Bio::Tools::Lucy - Object for analyzing the output from Lucy, a vector and quality trimming program from TIGR

SYNOPSIS

  # Create the Lucy object from an existing Lucy output file
  @params = ('seqfile' => 'lucy.seq', 'lucy_verbose' => 1);
  $lucyObj = Bio::Tools::Lucy->new(@params);

  # Get names of all sequences
  $names = $lucyObj->get_sequence_names();

  #  Print seq and qual values for sequences >400 bp in order to run CAP3
  foreach $name (@$names) {
      next unless $lucyObj->length_clear($name) > 400;
      print SEQ ">$name\n", $lucyObj->sequence($name), "\n";
      print QUAL ">$name\n", $lucyObj->quality($name), "\n";
  }

  # Get an array of Bio::PrimarySeq objects
  @seqObjs = $lucyObj->get_Seq_Objs();

DESCRIPTION

Bio::Tools::Lucy.pm provides methods for analyzing the sequence and quality values generated by Lucy program from TIGR.

Lucy will identify vector, poly-A/T tails, and poor quality regions in a sequence. (www.genomics.purdue.edu/gcg/other/lucy.pdf)

The input to Lucy can be the Phred sequence and quality files generated from running Phred on a set of chromatograms.

Lucy can be obtained (free of charge to academic users) from www.tigr.org/softlab

There are a few methods that will only be available if you make some minor changes to the source for Lucy and then recompile. The changes are in the 'lucy.c' file and there is a diff between the original and the modified file in the Appendix

Please contact the author of this module if you have any problems making these modifications.

You do not have to make these modifications to use this module.

Creating a Lucy object

  @params = ('seqfile' => 'lucy.seq', 'adv_stderr' => 1, 
             'fwd_desig' => '_F', 'rev_desig' => '_R');
  $lucyObj = Bio::Tools::Lucy->new(@params);

Using a Lucy object

  You should get an array with the sequence names in order to use
  accessor methods.  Note: The Lucy binary program will fail unless
  the sequence names provided as input are unique.

  $names_ref = $lucyObj->get_sequence_names();

  This code snippet will produce a Fasta format file with sequence
  lengths and %GC in the description line.

  foreach $name (@$names) {
      print FILE ">$name\t",
                 $lucyObj->length_clear($name), "\t",
                 $lucyObj->per_GC($name), "\n",
                 $lucyObj->sequence($name), "\n";
  }


  Print seq and qual values for sequences >400 bp in order to assemble
  them with CAP3 (or other assembler).

  foreach $name (@$names) {
      next unless $lucyObj->length_clear($name) > 400;
      print SEQ ">$name\n", $lucyObj->sequence($name), "\n";
      print QUAL ">$name\n", $lucyObj->quality($name), "\n";
  }

  Get all the sequences as Bio::PrimarySeq objects (eg., for use with
  Bio::Tools::Run::StandaloneBlast to perform BLAST).

  @seqObjs = $lucyObj->get_Seq_Objs();

  Or use only those sequences that are full length and have a Poly-A
  tail.

  foreach $name (@$names) {
      next unless ($lucyObj->full_length($name) and $lucy->polyA($name));
      push @seqObjs, $lucyObj->get_Seq_Obj($name);
  }


  Get the names of those sequences that were rejected by Lucy.

  $rejects_ref = $lucyObj->get_rejects();

  Print the names of the rejects and 1 letter code for reason they
  were rejected.

  foreach $key (sort keys %$rejects_ref) {
      print "$key:  ", $rejects_ref->{$key};
  }

  There is a lot of other information available about the sequences
  analyzed by Lucy (see APPENDIX).  This module can be used with the
  DBI module to store this sequence information in a database.

FEEDBACK

Mailing Lists

User feedback is an integral part of the evolution of this and other Bioperl modules. Send your comments and suggestions preferably to one of the Bioperl mailing lists. Your participation is much appreciated.

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

Support

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.

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 the web:

  https://github.com/bioperl/bioperl-live/issues

AUTHOR

Andrew G. Walsh paeruginosa@hotmail.com

APPENDIX

Methods available to Lucy objects are described below. Please note that any method beginning with an underscore is considered internal and should not be called directly.

new

Title : new
Usage : $lucyObj = Bio::Tools::Lucy->new(seqfile => lucy.seq, rev_desig => '_R',
fwd_desig => '_F')
Function: creates a Lucy object from Lucy analysis files
Returns : reference to Bio::Tools::Lucy object
Args : seqfile Fasta sequence file generated by Lucy
qualfile Quality values file generated by Lucy
infofile Info file created when Lucy is run with -debug
'infofile' option
stderrfile Standard error captured from Lucy when Lucy is run
with -info option and STDERR is directed to stderrfile
(ie. lucy ... 2> stderrfile).
Info in this file will include sequences dropped for low
quality. If you've modified Lucy source (see adv_stderr below),
it will also include info on which sequences were dropped because
they were vector, too short, had no insert, and whether a poly-A
tail was found (if Lucy was run with -cdna option).
lucy_verbose verbosity level (0-1).
fwd_desig The string used to determine whether sequence is a
forward read.
The parser will assume that this match will occus at the
end of the sequence name string.
rev_desig As above, for reverse reads.
adv_stderr Can be set to a true value (1). Will only work if
you have modified
the Lucy source code as outlined in DESCRIPTION and capture
the standard error from Lucy.

If you don't provide filenames for qualfile, infofile or stderrfile, the module will assume that .qual, .info, and .stderr are the file extensions and search in the same directory as the .seq file for these files.

For example, if you create a Lucy object with $lucyObj = Bio::Tools::Lucy->new(seqfile =>lucy.seq), the module will find lucy.qual, lucy.info and lucy.stderr.

You can omit any or all of the quality, info or stderr files, but you will not be able to use all of the object methods (see method documentation below).

_parse

 Title   :  _parse
 Usage   :  n/a (internal function)
 Function:  called by new() to parse Lucy output files
 Returns :  nothing
 Args    :  none

get_Seq_Objs

 Title   :  get_Seq_Objs
 Usage   :  $lucyObj->get_Seq_Objs()
 Function:  returns an array of references to Bio::PrimarySeq objects 
            where -id = 'sequence name' and -seq = 'sequence'

 Returns :  array of Bio::PrimarySeq objects
 Args    :  none

get_Seq_Obj

 Title   :  get_Seq_Obj
 Usage   :  $lucyObj->get_Seq_Obj($seqname)
 Function:  returns reference to a Bio::PrimarySeq object where -id = 'sequence name'
            and -seq = 'sequence'
 Returns :  reference to Bio::PrimarySeq object
 Args    :  name of a sequence

get_sequence_names

 Title   :  get_sequence_names
 Usage   :  $lucyObj->get_sequence_names
 Function:  returns reference to an array of names of the sequences analyzed by Lucy.
            These names are required for most of the accessor methods.  
            Note: The Lucy binary will fail unless sequence names are unique.
 Returns :  array reference
 Args    :  none

sequence

 Title   :  sequence
 Usage   :  $lucyObj->sequence($seqname)
 Function:  returns the DNA sequence of one of the sequences analyzed by Lucy.
 Returns :  string
 Args    :  name of a sequence

quality

 Title   :  quality
 Usage   :  $lucyObj->quality($seqname)
 Function:  returns the quality values of one of the sequences analyzed by Lucy.
            This method depends on the user having provided a quality file.
 Returns :  string
 Args    :  name of a sequence

avg_quality

 Title   :  avg_quality
 Usage   :  $lucyObj->avg_quality($seqname)
 Function:  returns the average quality value for one of the sequences analyzed by Lucy.
 Returns :  float
 Args    :  name of a sequence

direction

 Title   :  direction
 Usage   :  $lucyObj->direction($seqname)
 Function:  returns the direction for one of the sequences analyzed by Lucy
            providing that 'fwd_desig' or 'rev_desig' were set when the
            Lucy object was created.
            Strings returned are: 'F' for forward, 'R' for reverse.  
 Returns :  string 
 Args    :  name of a sequence

length_raw

 Title   :  length_raw
 Usage   :  $lucyObj->length_raw($seqname)
 Function:  returns the length of a DNA sequence prior to quality/ vector 
            trimming by Lucy.
 Returns :  integer
 Args    :  name of a sequence

length_clear

 Title   :  length_clear
 Usage   :  $lucyObj->length_clear($seqname)
 Function:  returns the length of a DNA sequence following quality/ vector   
            trimming by Lucy.
 Returns :  integer
 Args    :  name of a sequence

start_clear

 Title   :  start_clear
 Usage   :  $lucyObj->start_clear($seqname)
 Function:  returns the beginning position of good quality, vector free DNA sequence 
            determined by Lucy.
 Returns :  integer
 Args    :  name of a sequence

end_clear

 Title   :  end_clear
 Usage   :  $lucyObj->end_clear($seqname)
 Function:  returns the ending position of good quality, vector free DNA sequence
            determined by Lucy.
 Returns :  integer
 Args    :  name of a sequence

per_GC

 Title   :  per_GC
 Usage   :  $lucyObj->per_GC($seqname)
 Function:  returns the percente of the good quality, vector free DNA sequence
            determined by Lucy.
 Returns :  float
 Args    :  name of a sequence

full_length

 Title   :  full_length
 Usage   :  $lucyObj->full_length($seqname)
 Function:  returns the truth value for whether or not the sequence read was
            full length (ie. vector present on both ends of read).  This method
            depends on the user having provided the 'info' file (Lucy must be
            run with the -debug 'info_filename' option to get this file).
 Returns :  boolean 
 Args    :  name of a sequence

polyA

 Title   :  polyA
 Usage   :  $lucyObj->polyA($seqname)
 Function:  returns the truth value for whether or not a poly-A tail was detected
            and clipped by Lucy.  This method depends on the user having modified
            the source for Lucy as outlined in DESCRIPTION and invoking Lucy with
            the -cdna option and saving the standard error.
            Note, the final sequence will not show the poly-A/T region.
 Returns :  boolean
 Args    :  name of a sequence

get_rejects

 Title   :  get_rejects
 Usage   :  $lucyObj->get_rejects()
 Function:  returns a hash containing names of rejects and a 1 letter code for the 
            reason Lucy rejected the sequence.
            Q- rejected because of low quality values
            S- sequence was short
            V- sequence was vector 
            E- sequence was empty
            P- poly-A/T trimming caused sequence to be too short
            In order to get the rejects, you must provide a file with the standard
            error from Lucy.  You will only get the quality category rejects unless
            you have modified the source and recompiled Lucy as outlined in DESCRIPTION.
 Returns :  hash reference
 Args    :  none

Diff for Lucy source code

  352a353,354
  >       /* AGW added next line */
  >       fprintf(stderr, "Empty: %s\n", seqs[i].name);
  639a642,643
  >         /* AGW added next line */
  >         fprintf(stderr, "Short/ no insert: %s\n", seqs[i].name);
  678c682,686
  <     if (left) seqs[i].left+=left;
  ---
  >     if (left) {
  >       seqs[i].left+=left;
  >       /*  AGW added next line */
  >       fprintf(stderr, "%s has PolyA (left).\n", seqs[i].name);
  >     }
  681c689,693
  <     if (right) seqs[i].right-=right;
  ---
  >     if (right) {
  >       seqs[i].right-=right;
  >       /* AGW added next line */
  >       fprintf(stderr, "%s has PolyA (right).\n", seqs[i].name);
  >     }
  682a695,696
  >       /* AGW added next line */
  >       fprintf(stderr, "Dropped PolyA: %s\n", seqs[i].name); 
  734a749,750
  >       /* AGW added next line */
  >       fprintf(stderr, "Vector: %s\n", seqs[i].name);

This patch is to be applied to lucy.c from the lucy-1.19p release

 277a278,279
 >       /* AGW added next line */
 >       fprintf(stderr, "Short/ no insert: %s\n", seqs[i].name);
 588c590,592
 <     if ((seqs[i].len=bases)<=0)
 ---
 >     if ((seqs[i].len=bases)<=0) {
 >       /* AGW added next line */
 >       fprintf(stderr, "Empty: %s\n", seqs[i].name);
 589a594
 >     }
 893c898,902
 <       if (left) seqs[i].left+=left;
 ---
 >       if (left) {
 >         seqs[i].left+=left;
 >         /*  AGW added next line */
 >         fprintf(stderr, "%s has PolyA (left).\n", seqs[i].name);
 >       }
 896c905,909
 <       if (right) seqs[i].right-=right;
 ---
 >       if (right) {
 >         seqs[i].right-=right;
 >         /*  AGW added next line */
 >         fprintf(stderr, "%s has PolyA (right).\n", seqs[i].name);
 >         }
 898a912,913
 >         /* AGW added next line */
 >         fprintf(stderr, "Dropped PolyA: %s\n", seqs[i].name);
 949a965,966
 >         /* AGW added next line */
 >           fprintf(stderr, "Vector: %s\n", seqs[i].name);