X-Git-Url: http://woldlab.caltech.edu/gitweb/?a=blobdiff_plain;f=alg%2Fsequence.cpp;h=521496d3497b2b96d4786819438d170f6ac50ce4;hb=f3c0553a22b0e4ddc39ee45f51725352d92e97f1;hp=845a01617016dd17dfcf7e8c96bc55d0fa43acd8;hpb=26b245a4e58853cb6e463a1ccf9538d6dca8709f;p=mussa.git

diff --git a/alg/sequence.cpp b/alg/sequence.cpp
index 845a016..521496d 100644
--- a/alg/sequence.cpp
+++ b/alg/sequence.cpp
@@ -22,6 +22,7 @@
 //                           ---------- sequence.cc -----------
 //                        ----------------------------------------
 #include <boost/filesystem/fstream.hpp>
+#include <boost/filesystem/operations.hpp>
 namespace fs = boost::filesystem;
 
 #include <boost/spirit/core.hpp>
@@ -31,33 +32,16 @@ namespace fs = boost::filesystem;
 namespace spirit = boost::spirit;
 
 #include "alg/sequence.hpp"
+#include "io.hpp"
 #include "mussa_exceptions.hpp"
 
 #include <string>
+#include <stdexcept>
 #include <iostream>
 #include <sstream>
+#include <set>
 
-annot::annot() 
- : begin(0),
-   end(0),
-   type(""),
-   name("")
-{
-}
-
-annot::annot(int begin, int end, std::string type, std::string name)
- : begin(begin),
-   end(end),
-   type(type),
-   name(name)
-{
-}
-
-annot::~annot()
-{
-}
-
-bool operator==(const annot& left, const annot& right)
+bool operator==(const motif& left, const motif& right)
 {
   return ((left.begin== right.begin) and
           (left.end == right.end) and
@@ -65,8 +49,20 @@ bool operator==(const annot& left, const annot& right)
           (left.name == right.name));
 }
 
-motif::motif(int begin, std::string motif)
- : annot(begin, begin+motif.size(), "motif", motif),
+motif::motif() 
+ : begin(0),
+   end(0),
+   type("motif"),
+   name(""),
+   sequence("")
+{
+}
+
+motif::motif(int begin_, std::string motif)
+ : begin(begin_),
+   end(begin_+motif.size()),
+   type("motif"),
+   name(motif),
    sequence(motif)
 {
 }
@@ -75,18 +71,10 @@ motif::~motif()
 {
 }
 
-const std::string Sequence::dna_alphabet("AaCcGgTtNn\012\015");
-const std::string Sequence::rna_alphabet("AaCcGgNnUu\012\015");
-  //! this is the general iupac alphabet for nucleotides
-const std::string Sequence::nucleic_iupac_alphabet("AaCcGgTtUuRrYyMmKkSsWwBbDdHhVvNn\012\015");
-  //! the protein alphabet
-const std::string Sequence::protein_alphabet("AaCcDdEeFfGgHhIiKkLlMmNnPpQqRrSsTtVvWwYy\012\015");
-
-Sequence::Sequence()
-  : parent(0),
-    seq_start(0),
-    seq_count(0),
-    strand(UnknownStrand)
+Sequence::Sequence(AlphabetRef alphabet)
+  : seq(new SeqSpan("", alphabet, SeqSpan::PlusStrand)),
+    annotation_list(new SeqSpanRefList), 
+    motif_list(new MotifList)
 {
 }
 
@@ -94,83 +82,95 @@ Sequence::~Sequence()
 {
 }
 
-Sequence::Sequence(const char *seq)
-  : parent(0),
-    seq_start(0),
-    seq_count(0),
-    strand(UnknownStrand),
-    header(""),
-    species("")
+Sequence::Sequence(const char *seq, AlphabetRef alphabet_, SeqSpan::strand_type strand_)
+  : header(""),
+    species(""),
+    annotation_list(new SeqSpanRefList),
+    motif_list(new MotifList)
 {
-  set_filtered_sequence(seq);
+  set_filtered_sequence(seq, alphabet_, 0, npos, strand_);
 }
 
-Sequence::Sequence(const std::string& seq) 
-  : parent(0),
-    seq_start(0),
-    seq_count(0),
-    strand(UnknownStrand),
-    header(""),
-    species("")
+Sequence::Sequence(const std::string& seq, 
+                   AlphabetRef alphabet_,
+                   SeqSpan::strand_type strand_) 
+  : header(""),
+    species(""),
+    annotation_list(new SeqSpanRefList),
+    motif_list(new MotifList)
 {
-  set_filtered_sequence(seq);
+  set_filtered_sequence(seq, alphabet_, 0, seq.size(), strand_);
 }
 
 Sequence::Sequence(const Sequence& o)
-  : parent(o.parent),
-    seq(o.seq),
-    seq_start(o.seq_start),
-    seq_count(o.seq_count),
-    strand(o.strand),
+  : seq(o.seq),
     header(o.header),
     species(o.species),
-    annots(o.annots),
+    annotation_list(o.annotation_list),
     motif_list(o.motif_list)
 {
 }
 
+Sequence::Sequence(const Sequence* o)
+  : seq(o->seq),
+    header(o->header),
+    species(o->species),
+    annotation_list(o->annotation_list),
+    motif_list(o->motif_list)
+{
+}
+
+Sequence::Sequence(const SequenceRef o)
+  : seq(new SeqSpan(o->seq)),
+    header(o->header),
+    species(o->species),
+    annotation_list(new SeqSpanRefList),
+    motif_list(o->motif_list)
+{
+  // copy over the annotations in the other sequence ref, 
+  // attaching them to our current sequence ref
+  for(SeqSpanRefList::const_iterator annot_i = o->annotation_list->begin();
+      annot_i != o->annotation_list->end();
+      ++annot_i)
+  {
+    size_type annot_begin= (*annot_i)->start();
+    size_type annot_count = (*annot_i)->size();
+    
+    SeqSpanRef new_annot(seq->subseq(annot_begin, annot_count));
+    new_annot->setAnnotations((*annot_i)->annotations());
+    annotation_list->push_back(new_annot);
+  }  
+}
+
+Sequence::Sequence(const SeqSpanRef& seq_ref)
+  : seq(seq_ref),
+    header(""),
+    species(""),
+    annotation_list(new SeqSpanRefList),
+    motif_list(new MotifList)
+{
+}
+
 Sequence &Sequence::operator=(const Sequence& s)
 {
   if (this != &s) {
-    parent = s.parent;
     seq = s.seq;
-    seq_start = s.seq_start;
-    seq_count = s.seq_count;
-    strand = s.strand;
     header = s.header;
     species = s.species;
-    annots = s.annots;
+    annotation_list = s.annotation_list;
     motif_list = s.motif_list;
   }
   return *this;
 }
 
-static void multiplatform_getline(std::istream& in, std::string& line)
+void Sequence::load_fasta(fs::path file_path, int seq_num, int start_index, int end_index)
 {
-  line.clear();
-  char c;
-  in.get(c);
-  while(in.good() and !(c == '\012' or c == '\015') ) {
-    line.push_back(c);
-    in.get(c);
-  }
-  // if we have cr-lf eat it
-  c = in.peek();
-  if (c=='\012' or c == '\015') {
-    in.get();
-  }
+  load_fasta(file_path, reduced_nucleic_alphabet, seq_num, start_index, end_index);
 }
 
 //! load a fasta file into a sequence
-/*! 
- * \param file_path the location of the fasta file in the filesystem
- * \param seq_num which sequence in the file to load
- * \param start_index starting position in the fasta sequence, 0 for beginning
- * \param end_index ending position in the fasta sequence, 0 for end
- * \return error message, empty string if no error. (gag!)
- */
-void Sequence::load_fasta(fs::path file_path, int seq_num,
-                          int start_index, int end_index)
+void Sequence::load_fasta(fs::path file_path, AlphabetRef a, 
+                          int seq_num, int start_index, int end_index)
 {
   fs::fstream data_file;
   data_file.open(file_path, std::ios::in);
@@ -180,7 +180,7 @@ void Sequence::load_fasta(fs::path file_path, int seq_num,
     throw mussa_load_error("Sequence File: "+file_path.string()+" not found"); 
   } else {
     try {
-      load_fasta(data_file, seq_num, start_index, end_index);
+      load_fasta(data_file, a, seq_num, start_index, end_index);
     } catch(sequence_empty_error e) {
       // there doesn't appear to be any sequence
       // catch and rethrow to include the filename
@@ -194,20 +194,34 @@ void Sequence::load_fasta(fs::path file_path, int seq_num,
       errormsg << file_path.native_file_string()
                << " did not have any fasta sequences" << std::endl;
       throw sequence_empty_file_error(errormsg.str());
+    } catch(sequence_invalid_load_error e) {
+      std::ostringstream msg;
+      msg << file_path.native_file_string();
+      msg << " " << e.what();
+      throw sequence_invalid_load_error(msg.str());
     }
   }
 }
 
+void Sequence::load_fasta(std::istream& file, 
+                          int seq_num, int start_index, int end_index)
+{
+  load_fasta(file, reduced_nucleic_alphabet, seq_num, start_index, end_index);
+}
+
 void
-Sequence::load_fasta(std::iostream& data_file, int seq_num, 
+Sequence::load_fasta(std::istream& data_file, AlphabetRef a, 
+                     int seq_num, 
                      int start_index, int end_index)
 {
   std::string file_data_line;
   int header_counter = 0;
+  size_t line_counter = 0;
   bool read_seq = true;
   std::string rev_comp;
   std::string sequence_raw;
   std::string seq_tmp;      // holds sequence during basic filtering
+  const Alphabet &alpha = Alphabet::get_alphabet(a);
 
   if (seq_num == 0) {
     throw mussa_load_error("fasta sequence number is 1 based (can't be 0)");
@@ -217,6 +231,7 @@ Sequence::load_fasta(std::iostream& data_file, int seq_num,
   while ( (!data_file.eof()) && (header_counter < seq_num) )
   {
     multiplatform_getline(data_file, file_data_line);
+    ++line_counter;
     if (file_data_line.substr(0,1) == ">")
       header_counter++;
   }
@@ -228,9 +243,24 @@ Sequence::load_fasta(std::iostream& data_file, int seq_num,
 
     while ( !data_file.eof() && read_seq ) {
       multiplatform_getline(data_file,file_data_line);
+      ++line_counter;
       if (file_data_line.substr(0,1) == ">")
         read_seq = false;
-      else sequence_raw += file_data_line;
+      else {
+        for (std::string::const_iterator line_i = file_data_line.begin();
+             line_i != file_data_line.end();
+             ++line_i)
+         {
+           if(alpha.exists(*line_i)) {
+             sequence_raw += *line_i;
+           } else {
+            std::ostringstream msg;
+            msg << "Unrecognized characters in fasta sequence at line ";
+            msg << line_counter;
+            throw sequence_invalid_load_error(msg.str());
+           }
+         }
+      }
     }
 
     // Lastly, if subselection of the sequence was specified we keep cut out
@@ -244,67 +274,71 @@ Sequence::load_fasta(std::iostream& data_file, int seq_num,
       std::string msg("The selected sequence appears to be empty"); 
       throw sequence_empty_error(msg);
     }
-    set_filtered_sequence(sequence_raw, start_index, end_index-start_index);
+    set_filtered_sequence(sequence_raw, a, start_index, end_index-start_index, SeqSpan::PlusStrand);
   } else {
     std::string errormsg("There were no fasta sequences");
     throw sequence_empty_file_error(errormsg);
   }
 }
 
-void Sequence::set_filtered_sequence(const std::string &old_seq, 
+void Sequence::set_filtered_sequence(const std::string &in_seq,
+                                     AlphabetRef alphabet_,
                                      size_type start,
                                      size_type count,
-                                     strand_type strand_)
+                                     SeqSpan::strand_type strand_)
 {
-  char conversionTable[257];
-
   if ( count == npos)
-    count = old_seq.size() - start;
-  boost::shared_ptr<seq_string> new_seq(new seq_string);
-  new_seq->reserve(count);
-
-  // Make a conversion table
-
-  // everything we don't specify below will become 'N'
-  for(int table_i=0; table_i < 256; table_i++)
-  {
-    conversionTable[table_i] = 'N';
-  }
-  // add end of string character for printing out table for testing purposes
-  conversionTable[256] = '\0';
-
-  // we want these to map to themselves - ie not to change
-  conversionTable[(int)'A'] = 'A';
-  conversionTable[(int)'T'] = 'T';
-  conversionTable[(int)'G'] = 'G';
-  conversionTable[(int)'C'] = 'C';
-  // this is to upcase
-  conversionTable[(int)'a'] = 'A';
-  conversionTable[(int)'t'] = 'T';
-  conversionTable[(int)'g'] = 'G';
-  conversionTable[(int)'c'] = 'C';
+    count = in_seq.size() - start;
+  std::string new_seq;
+  new_seq.reserve(count);
 
   // finally, the actual conversion loop
-  for(std::string::size_type seq_index = 0; seq_index < count; seq_index++)
+  const Alphabet& alpha_impl = Alphabet::get_alphabet(alphabet_); // go get one of our actual alphabets
+  std::string::const_iterator seq_i = in_seq.begin()+start;
+  for(size_type i = 0; i != count; ++i, ++seq_i)
   {
-    new_seq->append(1, conversionTable[ (int)old_seq[seq_index+start]]);
+    if (alpha_impl.exists(*seq_i)) {
+      new_seq.append(1, toupper(*seq_i));
+    } else {
+      new_seq.append(1, 'N');
+    }
   }
-  parent = 0;
-  seq = new_seq;
-  seq_start = 0;
-  seq_count = count;
-  strand = strand_;
+  SeqSpanRef new_seq_ref(new SeqSpan(new_seq, alphabet_, strand_)); 
+  seq = new_seq_ref;
 }
 
 void
 Sequence::load_annot(fs::path file_path, int start_index, int end_index)
 {
+  if (not fs::exists(file_path)) {
+    throw mussa_load_error("Annotation File " + file_path.string() + " was not found");
+  }
+  if (fs::is_directory(file_path)) {
+    throw mussa_load_error(file_path.string() +
+            " is a directory, please provide a file for annotations."
+          );
+  }    
   fs::fstream data_stream(file_path, std::ios::in);
   if (!data_stream)
   {
-    throw mussa_load_error("Sequence File: " + file_path.string() + " not found");
+    throw mussa_load_error("Error loading annotation file " + file_path.string());
   }
 
+  try {  
+    load_annot(data_stream, start_index, end_index);
+  }  catch(annotation_load_error e) {
+    std::ostringstream msg;
+    msg << file_path.native_file_string()
+        << " "
+        << e.what();
+    throw annotation_load_error(msg.str());
+  }
+  data_stream.close();
+}
+
+void
+Sequence::load_annot(std::istream& data_stream, int start_index, int end_index)
+{
   // so i should probably be passing the parse function some iterators
   // but the annotations files are (currently) small, so i think i can 
   // get away with loading the whole file into memory
@@ -314,8 +348,7 @@ Sequence::load_annot(fs::path file_path, int start_index, int end_index)
     data_stream.get(c);
     data.push_back(c);
   }
-  data_stream.close();
-        
+
   parse_annot(data, start_index, end_index);
 }
 
@@ -337,30 +370,36 @@ Sequence::load_annot(fs::path file_path, int start_index, int end_index)
  */
   
 struct push_back_annot {
-  std::list<annot>& annot_list;
+  Sequence* parent;
+  SeqSpanRefListRef children;
   int& begin;
   int& end;
   std::string& name;
   std::string& type;
+  int &parsed;
 
-  push_back_annot(std::list<annot>& annot_list_, 
+  push_back_annot(Sequence* parent_seq,
+                  SeqSpanRefListRef children_list,
                   int& begin_, 
                   int& end_, 
                   std::string& name_, 
-                  std::string& type_) 
-  : annot_list(annot_list_), 
+                  std::string& type_,
+                  int &parsed_) 
+  : parent(parent_seq),
+    children(children_list),
     begin(begin_),
     end(end_),
     name(name_),
-    type(type_)
+    type(type_),
+    parsed(parsed_)
   {
   }
 
   void operator()(std::string::const_iterator, 
                   std::string::const_iterator) const 
   {
-    //std::cout << "adding annot: " << begin << "|" << end << "|" << name << "|" << type << std::endl;
-    annot_list.push_back(annot(begin, end, name, type));
+    children->push_back(parent->make_annotation(name, type, begin, end));
+    ++parsed;
   };
 };
 
@@ -368,13 +407,16 @@ struct push_back_seq {
   std::list<Sequence>& seq_list;
   std::string& name;
   std::string& seq;
+  int &parsed;
 
   push_back_seq(std::list<Sequence>& seq_list_,
                 std::string& name_, 
-                std::string& seq_)
+                std::string& seq_,
+                int &parsed_)
   : seq_list(seq_list_), 
     name(name_),
-    seq(seq_)
+    seq(seq_),
+    parsed(parsed_)
   {
   }
 
@@ -394,173 +436,178 @@ struct push_back_seq {
     Sequence s(new_seq);
     s.set_fasta_header(name);
     seq_list.push_back(s);
+    ++parsed;
   };
 };
 
-bool
+void
 Sequence::parse_annot(std::string data, int start_index, int end_index)
 {
   int start=0;
   int end=0;
   std::string name;
   std::string type;
-  std::string seq;
+  std::string seqstr;
+  SeqSpanRefListRef parsed_annots(new SeqSpanRefList);
   std::list<Sequence> query_seqs;
-
-  bool status = spirit::parse(data.begin(), data.end(),
-                (
-                 //begin grammar
-                   !(
-                      (
-                        spirit::alpha_p >> 
-                        +(spirit::graph_p)
-                      )[spirit::assign_a(species)] >> 
-                      +(spirit::space_p)
-                    ) >>
-                    *(
-                       ( // ignore html tags
-                         *(spirit::space_p) >>
-                         spirit::ch_p('<') >> 
-                         +(~spirit::ch_p('>')) >>
-                         spirit::ch_p('>') >>
-                         *(spirit::space_p)
-                       )
-                     |
-                      ( // parse an absolute location name
-                       (spirit::uint_p[spirit::assign_a(start)] >> 
-                        +spirit::space_p >>
-                        spirit::uint_p[spirit::assign_a(end)] >> 
-                        +spirit::space_p >>
-                        ( 
-                           spirit::alpha_p >> 
-                           *spirit::graph_p
-                        )[spirit::assign_a(name)] >> 
-                        // optional type
-                        !(
-                            +spirit::space_p >>
-                            (
-                              spirit::alpha_p >>
-                              *spirit::graph_p
-                            )[spirit::assign_a(type)]
-                        )
-                        // to understand how this group gets set
-                        // read the comment above struct push_back_annot
-                       )[push_back_annot(annots, start, end, type, name)]
-                     |
-                      ((spirit::ch_p('>')|spirit::str_p("&gt;")) >> 
-                         (*(spirit::print_p))[spirit::assign_a(name)] >>
-                         spirit::eol_p >> 
-                         (+(spirit::chset<>(nucleic_iupac_alphabet.c_str())))[spirit::assign_a(seq)]
-                       )[push_back_seq(query_seqs, name, seq)]
-                      ) >>
-                      *spirit::space_p
+  int parsed=0;
+
+  bool ok = spirit::parse(data.begin(), data.end(),
+              (
+               //begin grammar
+                 !(
+                    (
+                      spirit::alpha_p >> 
+                      +(spirit::graph_p)
+                    )[spirit::assign_a(species)] >> 
+                    +(spirit::space_p)
+                  ) >>
+                  *(
+                     ( // ignore html tags
+                       *(spirit::space_p) >>
+                       spirit::ch_p('<') >> 
+                       +(~spirit::ch_p('>')) >>
+                       spirit::ch_p('>') >>
+                       *(spirit::space_p)
                      )
-                //end grammar
-                )).full;
-                
-  // go seearch for query sequences 
+                   |
+                    ( // parse an absolute location name
+                     (spirit::uint_p[spirit::assign_a(start)] >> 
+                      +spirit::space_p >>
+                      spirit::uint_p[spirit::assign_a(end)] >> 
+                      +spirit::space_p >>
+                      ( 
+                         spirit::alpha_p >> 
+                         *spirit::graph_p
+                      )[spirit::assign_a(name)] >> 
+                      // optional type
+                      !(
+                          +spirit::space_p >>
+                          (
+                            spirit::alpha_p >>
+                            *spirit::graph_p
+                          )[spirit::assign_a(type)]
+                      )
+                      // to understand how this group gets set
+                      // read the comment above struct push_back_annot
+                     )[push_back_annot(this, parsed_annots, start, end, name, type, parsed)]
+                   |
+                    ((spirit::ch_p('>')|spirit::str_p("&gt;")) >> 
+                       (*(spirit::print_p))[spirit::assign_a(name)] >>
+                       spirit::eol_p >> 
+                       (+(spirit::chset<>(Alphabet::nucleic_cstr)))[spirit::assign_a(seqstr)]
+                     )[push_back_seq(query_seqs, name, seqstr, parsed)]
+                    ) >>
+                    *spirit::space_p
+                   )
+              //end grammar
+              )).full;
+  if (not ok) {
+    std::stringstream msg;
+    msg << "Error parsing annotation #" << parsed;
+    throw annotation_load_error(msg.str());
+  }
+  // If everything loaded correctly add the sequences to our annotation list
+  // add newly parsed annotations to our sequence
+  std::copy(parsed_annots->begin(), parsed_annots->end(), std::back_inserter(*annotation_list));  
+  // go search for query sequences 
   find_sequences(query_seqs.begin(), query_seqs.end());
-  return status;
 }
 
-void Sequence::add_annotation(const annot& a)
+void Sequence::add_annotation(const SeqSpanRef a)
 {
-  annots.push_back(a);
+  annotation_list->push_back(a);
 }
 
-const std::list<annot>& Sequence::annotations() const
+void Sequence::add_annotation(std::string name, std::string type, size_type start, size_type stop)
 {
-  return annots;
+  add_annotation(make_annotation(name, type, start, stop));
 }
 
-Sequence
-Sequence::subseq(int start, int count)
+SeqSpanRef 
+Sequence::make_annotation(std::string name, std::string type, size_type start, size_type stop) const
 {
-  if (!seq) {
-    Sequence new_seq;
-    return new_seq;
+  // we want things to be in the positive direction
+  if (stop < start) {
+    size_type tmp = start;
+    start = stop;
+    stop = tmp;
   }
+  size_type count = stop - start;
+  SeqSpanRef new_annot(seq->subseq(start, count,  SeqSpan::UnknownStrand));
+  AnnotationsRef metadata(new Annotations(name));
+  metadata->set("type", type);
+  new_annot->setAnnotations(metadata);
+  return new_annot;
+}
 
-  // there might be an off by one error with start+count > size()
-  if ( count == npos || start+count > size()) {
-    count = size()-start;
-  }
-  Sequence new_seq(*this);
-  new_seq.parent = this;
-  new_seq.seq_start = seq_start+start;
-  new_seq.seq_count = count;
+const SeqSpanRefList& Sequence::annotations() const
+{
+  return *annotation_list;
+}
 
+void Sequence::copy_children(Sequence &new_seq, size_type start, size_type count) const
+{
   new_seq.motif_list = motif_list;
-  new_seq.annots.clear();
-  // attempt to copy & reannotate position based annotations 
-  int end = start+count;
+  new_seq.annotation_list.reset(new SeqSpanRefList);
 
-  for(std::list<annot>::const_iterator annot_i = annots.begin();
-      annot_i != annots.end();
+  for(SeqSpanRefList::const_iterator annot_i = annotation_list->begin();
+      annot_i != annotation_list->end();
       ++annot_i)
   {
-    int annot_begin= annot_i->begin;
-    int annot_end = annot_i->end;
+    size_type annot_begin= (*annot_i)->start();
+    size_type annot_end = (*annot_i)->stop();
 
-    if (annot_begin < end) {
+    if (annot_begin < start+count) {
       if (annot_begin >= start) {
         annot_begin -= start;
       } else {
         annot_begin = 0;
       }
 
-      if (annot_end < end) {
+      if (annot_end < start+count) {
         annot_end -= start;
       } else {
         annot_end = count;
       }
-
-      annot new_annot(annot_begin, annot_end, annot_i->type, annot_i->name);
-      new_seq.annots.push_back(new_annot);
+      SeqSpanRef new_annot(new_seq.seq->subseq(annot_begin, annot_end));
+      new_annot->setAnnotations((*annot_i)->annotations());
+      new_seq.annotation_list->push_back(new_annot);
     }
   }
-
-  return new_seq;
 }
 
-std::string
-Sequence::rev_comp() const
+Sequence
+Sequence::subseq(size_type start, size_type count, SeqSpan::strand_type strand) const
 {
-  std::string rev_comp;
-  char conversionTable[257];
-  int seq_i, table_i, len;
-
-  len = length();
-  rev_comp.reserve(len);
-  // make a conversion table
-  // init all parts of conversion table to '~' character
-  // '~' I doubt will ever appear in a sequence file (jeez, I hope)
-  // and may the fleas of 1000 camels infest the genitals of any biologist (and
-  // seven generations of their progeny) who decides to make it mean
-  // something special!!!
-  // PS - double the curse for any smartass non-biologist who tries it as well
-  for(table_i=0; table_i < 256; table_i++)
-  {
-    conversionTable[table_i] = '~';
+  // FIXME: should i really allow a subsequence of an empty sequence?
+  if (!seq) {
+    Sequence new_seq;
+    return new_seq;
   }
-  // add end of string character for printing out table for testing purposes
-  conversionTable[256] = '\0';
-
-  // add in the characters for the bases we want to convert
-  conversionTable[(int)'A'] = 'T';
-  conversionTable[(int)'T'] = 'A';
-  conversionTable[(int)'G'] = 'C';
-  conversionTable[(int)'C'] = 'G';
-  conversionTable[(int)'N'] = 'N';
 
-  // finally, the actual conversion loop
-  for(seq_i = len - 1; seq_i >= 0; seq_i--)
-  {
-    table_i = (int) at(seq_i);
-    rev_comp += conversionTable[table_i];
+  Sequence new_seq(*this);
+  new_seq.seq = seq->subseq(start, count, strand);
+  if (seq->annotations()) {
+    AnnotationsRef a(new Annotations(*(seq->annotations())));
+    new_seq.seq->setAnnotations(a);
   }
+  copy_children(new_seq, start, count);
+  
+  return new_seq;
+}
+
 
-  return rev_comp;
+// FIXME: This needs to be moved into SeqSpan
+Sequence Sequence::rev_comp() const
+{
+  // a reverse complement is the whole opposite strand 
+  return subseq(0, npos, SeqSpan::OppositeStrand);
+}
+
+const Alphabet& Sequence::get_alphabet() const
+{
+  return (seq) ? seq->get_alphabet() : Alphabet::empty_alphabet(); 
 }
 
 void Sequence::set_fasta_header(std::string header_)
@@ -596,17 +643,14 @@ Sequence::get_name() const
     return "";
 }
 
-void Sequence::set_sequence(const std::string& s) 
+void Sequence::set_sequence(const std::string& s, AlphabetRef a) 
 {
-  set_filtered_sequence(s);
+  set_filtered_sequence(s, a, 0, s.size(), SeqSpan::PlusStrand);
 }
 
 std::string Sequence::get_sequence() const
 {
-  if (seq) 
-    return *seq;
-  else
-    return std::string();
+  return seq->sequence();
 }
 
 Sequence::const_reference Sequence::operator[](Sequence::size_type i) const
@@ -614,84 +658,24 @@ Sequence::const_reference Sequence::operator[](Sequence::size_type i) const
   return at(i);
 }
 
-Sequence::const_reference Sequence::at(Sequence::size_type i) const
-{
-  if (!seq) throw std::out_of_range("empty sequence");
-  return seq->at(i+seq_start);
-}
-
 void
 Sequence::clear()
 {
-  parent = 0;
   seq.reset();
-  seq_start = 0;
-  seq_count = 0;
-  strand = UnknownStrand;
   header.clear();
   species.clear();
-  annots.clear();
-  motif_list.clear();
-}
-
-const char *Sequence::c_str() const
-{
-  if (seq) 
-    return seq->c_str()+seq_start;
-  else 
-    return 0;
-}
-
-Sequence::const_iterator Sequence::begin() const
-{
-  if (seq and seq_count != 0)
-    return seq->begin()+seq_start;
-  else 
-    return Sequence::const_iterator(0);
-}
-
-Sequence::const_iterator Sequence::end() const
-{
-  if (seq and seq_count != 0) {
-    return seq->begin() + seq_start + seq_count;
-  } else {
-    return Sequence::const_iterator(0);
-  }
-}
-
-bool Sequence::empty() const
-{
-  return (seq_count == 0) ? true : false;
-}
-
-Sequence::size_type Sequence::start() const
-{
-  if (parent)
-    return seq_start - parent->start();
-  else
-    return seq_start;
-}
-
-Sequence::size_type Sequence::stop() const
-{
-  return start() + seq_count;
-}
-
-Sequence::size_type Sequence::size() const
-{
-  return seq_count;
-}
-
-Sequence::size_type Sequence::length() const
-{
-  return size();
+  annotation_list.reset(new SeqSpanRefList);
+  motif_list.reset(new MotifList);
 }
 
 void
 Sequence::save(fs::fstream &save_file)
 {
+  std::string type("type");
+  std::string empty_str("");
   //fstream save_file;
-  std::list<annot>::iterator annots_i;
+  SeqSpanRefList::iterator annots_i;
+  AnnotationsRef metadata;
 
   // not sure why, or if i'm doing something wrong, but can't seem to pass
   // file pointers down to this method from the mussa control class
@@ -704,45 +688,149 @@ Sequence::save(fs::fstream &save_file)
 
   save_file << "<Annotations>" << std::endl;
   save_file << species << std::endl;
-  for (annots_i = annots.begin(); annots_i != annots.end(); ++annots_i)
+  for (annots_i = annotation_list->begin(); 
+       annots_i != annotation_list->end(); 
+       ++annots_i)
   {
-    save_file << annots_i->begin << " " << annots_i->end << " " ;
-    save_file << annots_i->name << " " << annots_i->type << std::endl;
+    metadata = (*annots_i)->annotations();
+    save_file << (*annots_i)->parentStart() << " " << (*annots_i)->parentStop() << " " ;
+    save_file << metadata->name() << " " 
+              << metadata->getdefault(type, empty_str) << std::endl;
   }
   save_file << "</Annotations>" << std::endl;
   //save_file.close();
 }
 
-void
-Sequence::load_museq(fs::path load_file_path, int seq_num)
+//void
+//Sequence::load_museq(fs::path load_file_path, int seq_num)
+//{
+//  fs::fstream load_file;
+//  std::string file_data_line;
+//  int seq_counter;
+//  //annot an_annot;
+//  int annot_begin;
+//  int annot_end;
+//  std::string annot_name;
+//  std::string annot_type;
+//  
+//  std::string::size_type space_split_i;
+//  std::string annot_value;
+//
+//  annotation_list.reset(new SeqSpanRefList);
+//  
+//  load_file.open(load_file_path, std::ios::in);
+//
+//  seq_counter = 0;
+//  // search for the seq_num-th sequence 
+//  while ( (!load_file.eof()) && (seq_counter < seq_num) )
+//  {
+//    getline(load_file,file_data_line);
+//    if (file_data_line == "<Sequence>")
+//      seq_counter++;
+//  }
+//  getline(load_file, file_data_line);
+//  // looks like the sequence is written as a single line
+//  set_filtered_sequence(file_data_line, reduced_dna_alphabet, 0, file_data_line.size(), SeqSpan::PlusStrand);
+//  getline(load_file, file_data_line);
+//  getline(load_file, file_data_line);
+//  if (file_data_line == "<Annotations>")
+//  {
+//    getline(load_file, file_data_line);
+//    species = file_data_line;
+//    while ( (!load_file.eof())  && (file_data_line != "</Annotations>") )
+//    {
+//      getline(load_file,file_data_line);
+//      if ((file_data_line != "") && (file_data_line != "</Annotations>"))  
+//      {
+//        // need to get 4 values...almost same code 4 times...
+//        // get annot start index
+//        space_split_i = file_data_line.find(" ");
+//        annot_value = file_data_line.substr(0,space_split_i);
+//        annot_begin = atoi (annot_value.c_str());
+//        file_data_line = file_data_line.substr(space_split_i+1);
+//        // get annot end index
+//        space_split_i = file_data_line.find(" ");
+//        annot_value = file_data_line.substr(0,space_split_i);
+//        annot_end = atoi (annot_value.c_str());
+//
+//        if (space_split_i == std::string::npos)  // no entry for type or name
+//        {
+//          std::cout << "seq, annots - no type or name\n";
+//          annot_name = "";
+//          annot_type = "";
+//        }
+//        else   // else get annot type
+//        {
+//          file_data_line = file_data_line.substr(space_split_i+1);
+//          space_split_i = file_data_line.find(" ");
+//          annot_value = file_data_line.substr(0,space_split_i);
+//          //an_annot.type = annot_value;
+//          annot_type = annot_value;
+//          if (space_split_i == std::string::npos)  // no entry for name
+//          {
+//            std::cout << "seq, annots - no name\n";
+//            annot_name = "";
+//          }
+//          else          // get annot name
+//          {
+//            file_data_line = file_data_line.substr(space_split_i+1);
+//            space_split_i = file_data_line.find(" ");
+//            annot_value = file_data_line.substr(0,space_split_i);
+//            // this seems like its wrong?
+//            annot_type = annot_value;
+//          }
+//        }
+//        add_annotation(annot_name, annot_type, annot_begin, annot_end);
+//      }
+//      //std::cout << "seq, annots: " << an_annot.start << ", " << an_annot.end
+//      //     << "-->" << an_annot.type << "::" << an_annot.name << std::endl;
+//    }
+//  }
+//  load_file.close();
+//}
+
+SequenceRef Sequence::load_museq(boost::filesystem::fstream& load_file)
 {
-  fs::fstream load_file;
+  boost::shared_ptr<Sequence> seq(new Sequence);
   std::string file_data_line;
   int seq_counter;
-  annot an_annot;
+  //annot an_annot;
+  int annot_begin;
+  int annot_end;
+  std::string annot_name;
+  std::string annot_type;
+  
   std::string::size_type space_split_i;
   std::string annot_value;
 
-  annots.clear();
-  load_file.open(load_file_path, std::ios::in);
+  //seq->annotation_list.reset(new SeqSpanRefList);
 
   seq_counter = 0;
-  // search for the seq_num-th sequence 
+  // search for the next sequence
+  int seq_num = 1;
   while ( (!load_file.eof()) && (seq_counter < seq_num) )
   {
     getline(load_file,file_data_line);
     if (file_data_line == "<Sequence>")
       seq_counter++;
   }
+  
+  // Could not find next sequence
+  if (load_file.eof())
+  {
+    seq.reset();
+    return seq;
+  }
+  
   getline(load_file, file_data_line);
   // looks like the sequence is written as a single line
-  set_filtered_sequence(file_data_line);
+  seq->set_filtered_sequence(file_data_line, reduced_dna_alphabet, 0, file_data_line.size(), SeqSpan::PlusStrand);
   getline(load_file, file_data_line);
   getline(load_file, file_data_line);
   if (file_data_line == "<Annotations>")
   {
     getline(load_file, file_data_line);
-    species = file_data_line;
+    seq->set_species(file_data_line);
     while ( (!load_file.eof())  && (file_data_line != "</Annotations>") )
     {
       getline(load_file,file_data_line);
@@ -752,148 +840,50 @@ Sequence::load_museq(fs::path load_file_path, int seq_num)
         // get annot start index
         space_split_i = file_data_line.find(" ");
         annot_value = file_data_line.substr(0,space_split_i);
-        an_annot.begin = atoi (annot_value.c_str());
+        annot_begin = atoi (annot_value.c_str());
         file_data_line = file_data_line.substr(space_split_i+1);
         // get annot end index
         space_split_i = file_data_line.find(" ");
         annot_value = file_data_line.substr(0,space_split_i);
-        an_annot.end = atoi (annot_value.c_str());
+        annot_end = atoi (annot_value.c_str());
 
         if (space_split_i == std::string::npos)  // no entry for type or name
         {
           std::cout << "seq, annots - no type or name\n";
-          an_annot.type = "";
-          an_annot.name = "";
+          annot_name = "";
+          annot_type = "";
         }
         else   // else get annot type
         {
           file_data_line = file_data_line.substr(space_split_i+1);
           space_split_i = file_data_line.find(" ");
           annot_value = file_data_line.substr(0,space_split_i);
-          an_annot.type = annot_value;
+          //an_annot.type = annot_value;
+          annot_type = annot_value;
           if (space_split_i == std::string::npos)  // no entry for name
           {
             std::cout << "seq, annots - no name\n";
-            an_annot.name = "";
+            annot_name = "";
           }
           else          // get annot name
           {
             file_data_line = file_data_line.substr(space_split_i+1);
             space_split_i = file_data_line.find(" ");
             annot_value = file_data_line.substr(0,space_split_i);
-            an_annot.type = annot_value;
+            // this seems like its wrong?
+            annot_type = annot_value;
           }
         }
-        annots.push_back(an_annot);  // don't forget to actually add the annot
+        seq->add_annotation(annot_name, annot_type, annot_begin, annot_end);
       }
       //std::cout << "seq, annots: " << an_annot.start << ", " << an_annot.end
       //     << "-->" << an_annot.type << "::" << an_annot.name << std::endl;
     }
   }
-  load_file.close();
-}
-
-std::string
-Sequence::rc_motif(std::string a_motif) const
-{
-  std::string rev_comp;
-  char conversionTable[257];
-  int seq_i, table_i, len;
-
-  len = a_motif.length();
-  rev_comp.reserve(len);
-
-  for(table_i=0; table_i < 256; table_i++)
-  {
-    conversionTable[table_i] = '~';
-  }
-  // add end of std::string character for printing out table for testing purposes
-  conversionTable[256] = '\0';
-
-  // add in the characters for the bases we want to convert (IUPAC)
-  conversionTable[(int)'A'] = 'T';
-  conversionTable[(int)'T'] = 'A';
-  conversionTable[(int)'G'] = 'C';
-  conversionTable[(int)'C'] = 'G';
-  conversionTable[(int)'N'] = 'N';
-  conversionTable[(int)'M'] = 'K';
-  conversionTable[(int)'R'] = 'Y';
-  conversionTable[(int)'W'] = 'W';
-  conversionTable[(int)'S'] = 'S';
-  conversionTable[(int)'Y'] = 'R';
-  conversionTable[(int)'K'] = 'M';
-  conversionTable[(int)'V'] = 'B';
-  conversionTable[(int)'H'] = 'D';
-  conversionTable[(int)'D'] = 'H';
-  conversionTable[(int)'B'] = 'V';
-
-  // finally, the actual conversion loop
-  for(seq_i = len - 1; seq_i >= 0; seq_i--)
-  {
-    //std::cout << "** i = " << seq_i << " bp = " << 
-    table_i = (int) a_motif[seq_i];
-    rev_comp += conversionTable[table_i];
-  }
-
-  //std::cout << "seq: " << a_motif << std::endl;
-  //std::cout << "rc:  " << rev_comp << std::endl;
-
-  return rev_comp;
+  //load_file.close();
+  return seq;
 }
 
-std::string
-Sequence::motif_normalize(const std::string& a_motif)
-{
-  std::string valid_motif;
-  int seq_i, len;
-
-  len = a_motif.length();
-  valid_motif.reserve(len);
-
-  // this just upcases IUPAC symbols.  Eventually should return an error if non IUPAC is present.
-  // current nonIUPAC symbols are omitted, which is not reported atm
-  for(seq_i = 0; seq_i < len; seq_i++)
-  {
-    if ((a_motif[seq_i] == 'a') || (a_motif[seq_i] == 'A'))
-      valid_motif += 'A';
-    else if ((a_motif[seq_i] == 't') || (a_motif[seq_i] == 'T'))
-      valid_motif += 'T';
-    else if ((a_motif[seq_i] == 'g') || (a_motif[seq_i] == 'G'))
-      valid_motif += 'G';
-    else if ((a_motif[seq_i] == 'c') || (a_motif[seq_i] == 'C'))
-      valid_motif += 'C';
-    else if ((a_motif[seq_i] == 'n') || (a_motif[seq_i] == 'N'))
-      valid_motif += 'N';
-    else if ((a_motif[seq_i] == 'm') || (a_motif[seq_i] == 'M'))
-      valid_motif += 'M';
-    else if ((a_motif[seq_i] == 'r') || (a_motif[seq_i] == 'R'))
-      valid_motif += 'R';
-    else if ((a_motif[seq_i] == 'w') || (a_motif[seq_i] == 'W'))
-      valid_motif += 'W';
-    else if ((a_motif[seq_i] == 's') || (a_motif[seq_i] == 'S'))
-      valid_motif += 'S';
-    else if ((a_motif[seq_i] == 'y') || (a_motif[seq_i] == 'Y'))
-      valid_motif += 'Y';
-    else if ((a_motif[seq_i] == 'k') || (a_motif[seq_i] == 'K'))
-      valid_motif += 'G';
-    else if ((a_motif[seq_i] == 'v') || (a_motif[seq_i] == 'V'))
-      valid_motif += 'V';
-    else if ((a_motif[seq_i] == 'h') || (a_motif[seq_i] == 'H'))
-      valid_motif += 'H';
-    else if ((a_motif[seq_i] == 'd') || (a_motif[seq_i] == 'D'))
-      valid_motif += 'D';
-    else if ((a_motif[seq_i] == 'b') || (a_motif[seq_i] == 'B'))
-      valid_motif += 'B';
-    else {
-      std::string msg = "Letter ";
-      msg += a_motif[seq_i];
-      msg += " is not a valid IUPAC symbol";
-      throw motif_normalize_error(msg);
-    }
-  }
-  //std::cout << "valid_motif is: " << valid_motif << std::endl;
-  return valid_motif;
-}
 
 void Sequence::add_motif(const Sequence& a_motif)
 {
@@ -903,122 +893,96 @@ void Sequence::add_motif(const Sequence& a_motif)
       motif_start_i != motif_starts.end();
       ++motif_start_i)
   {
-    motif_list.push_back(motif(*motif_start_i, a_motif.get_sequence()));
+    motif_list->push_back(motif(*motif_start_i, a_motif.get_sequence()));
   }
 }
 
 void Sequence::clear_motifs()
 {
-  motif_list.clear();
+  if (motif_list) 
+    motif_list->clear();
+  else 
+    motif_list.reset(new MotifList);
 }
 
-const std::list<motif>& Sequence::motifs() const
+const Sequence::MotifList& Sequence::motifs() const
 {
-  return motif_list;
+  return *motif_list;
 }
 
 std::vector<int>
-Sequence::find_motif(const std::string& a_motif) const
+Sequence::find_motif(const Sequence& a_motif) const
 {
   std::vector<int> motif_match_starts;
-  std::string norm_motif_rc;
+  Sequence norm_motif_rc;
 
   motif_match_starts.clear();
+  // std::cout << "motif is: " << norm_motif << std::endl;
 
-  //std::cout << "motif is: " << a_motif << std::endl;
-  std::string norm_motif = motif_normalize(a_motif);
-  //std::cout << "motif is: " << a_motif << std::endl;
-
-  if (norm_motif.size() > 0)
+  if (a_motif.size() > 0)
   {
     //std::cout << "Sequence: none blank motif\n";
-    motif_scan(norm_motif, &motif_match_starts);
+    motif_scan(a_motif, &motif_match_starts);
 
-    norm_motif_rc = rc_motif(a_motif);
+    norm_motif_rc = a_motif.rev_comp();;
     // make sure not to do search again if it is a palindrome
-    if (norm_motif_rc != norm_motif) {
+    if (norm_motif_rc != a_motif) {
       motif_scan(norm_motif_rc, &motif_match_starts);
     }
   }
   return motif_match_starts;
 }
 
-std::vector<int>
-Sequence::find_motif(const Sequence& a_motif) const
-{
-  return find_motif(a_motif.get_sequence());
-}
-
 void
-Sequence::motif_scan(std::string a_motif, std::vector<int> * motif_match_starts) const
+Sequence::motif_scan(const Sequence& a_motif, std::vector<int> * motif_match_starts) const
 {
   // if there's no sequence we can't scan for it?
   // should this throw an exception?
   if (!seq) return;
 
-  std::string::const_iterator seq_c = seq->begin();
-  std::string::size_type seq_i;
-  int motif_i, motif_len;
+  std::string::size_type seq_i = 0;
+  Sequence::size_type motif_i = 0;
+  Sequence::size_type motif_len = a_motif.length();
+  Sequence::value_type motif_char;
+  Sequence::value_type seq_char;
 
-  //std::cout << "Sequence: motif, seq len = " << sequence.length() << std::endl; 
-  motif_len = a_motif.length();
-
-  //std::cout << "motif_length: " << motif_len << std::endl;
-  //std::cout << "RAAARRRRR\n";
-
-  motif_i = 0;
-
-  //std::cout << "motif: " << a_motif << std::endl;
-
-  //std::cout << "Sequence: motif, length= " << length << std::endl;
-  seq_i = 0;
-  while (seq_i < length())
+  while (seq_i < size())
   {
-    //std::cout << seq_c[seq_i];
-    //std::cout << seq_c[seq_i] << "?" << a_motif[motif_i] << ":" << motif_i << " ";
     // this is pretty much a straight translation of Nora's python code
     // to match iupac letter codes
-    if (a_motif[motif_i] =='N')
+    motif_char = toupper(a_motif[motif_i]);
+    seq_char = toupper(seq->at(seq_i));
+    if (motif_char =='N')
       motif_i++;
-    else if (a_motif[motif_i] == seq_c[seq_i])
+    else if (motif_char == seq_char)
       motif_i++;
-    else if ((a_motif[motif_i] =='M') && 
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='C')))
+    else if ((motif_char =='M') && ((seq_char=='A') || (seq_char=='C')))
       motif_i++;
-    else if ((a_motif[motif_i] =='R') && 
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='G')))
+    else if ((motif_char =='R') && ((seq_char=='A') || (seq_char=='G')))
       motif_i++;
-    else if ((a_motif[motif_i] =='W') && 
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='T')))
+    else if ((motif_char =='W') && ((seq_char=='A') || (seq_char=='T')))
       motif_i++;
-    else if ((a_motif[motif_i] =='S') && 
-             ((seq_c[seq_i]=='C') || (seq_c[seq_i]=='G')))
+    else if ((motif_char =='S') && ((seq_char=='C') || (seq_char=='G')))
       motif_i++;
-    else if ((a_motif[motif_i] =='Y') && 
-             ((seq_c[seq_i]=='C') || (seq_c[seq_i]=='T')))
+    else if ((motif_char =='Y') && ((seq_char=='C') || (seq_char=='T')))
       motif_i++;
-    else if ((a_motif[motif_i] =='K') && 
-             ((seq_c[seq_i]=='G') || (seq_c[seq_i]=='T')))
+    else if ((motif_char =='K') && ((seq_char=='G') || (seq_char=='T')))
       motif_i++;
-    else if ((a_motif[motif_i] =='V') && 
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='C') ||
-              (seq_c[seq_i]=='G')))
+    else if ((motif_char =='V') && 
+             ((seq_char=='A') || (seq_char=='C') || (seq_char=='G')))
       motif_i++;
-    else if ((a_motif[seq_i] =='H') && 
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='C') ||
-              (seq_c[seq_i]=='T')))
+    else if ((motif_char =='H') && 
+             ((seq_char=='A') || (seq_char=='C') || (seq_char=='T')))
       motif_i++;
-    else if ((a_motif[motif_i] =='D') &&
-             ((seq_c[seq_i]=='A') || (seq_c[seq_i]=='G') ||
-              (seq_c[seq_i]=='T')))
+    else if ((motif_char =='D') &&
+             ((seq_char=='A') || (seq_char=='G') || (seq_char=='T')))
       motif_i++;
-    else if ((a_motif[motif_i] =='B') &&
-             ((seq_c[seq_i]=='C') || (seq_c[seq_i]=='G') ||
-              (seq_c[seq_i]=='T')))
+    else if ((motif_char =='B') &&
+             ((seq_char=='C') || (seq_char=='G') || (seq_char=='T')))
       motif_i++;
-
     else
     {
+      // if a motif doesn't match, erase our current trial and try again
       seq_i -= motif_i;
       motif_i = 0;
     }
@@ -1026,8 +990,6 @@ Sequence::motif_scan(std::string a_motif, std::vector<int> * motif_match_starts)
     // end Nora stuff, now we see if a match is found this pass
     if (motif_i == motif_len)
     {
-      //std::cout << "!!";
-      annot new_motif;
       motif_match_starts->push_back(seq_i - motif_len + 1);
       motif_i = 0;
     }
@@ -1042,16 +1004,11 @@ void Sequence::add_string_annotation(std::string a_seq,
 {
   std::vector<int> seq_starts = find_motif(a_seq);
   
-  //std::cout << "searching for " << a_seq << " found " << seq_starts.size() << std::endl;
-
   for(std::vector<int>::iterator seq_start_i = seq_starts.begin();
       seq_start_i != seq_starts.end();
       ++seq_start_i)
   {
-    annots.push_back(annot(*seq_start_i, 
-                           *seq_start_i+a_seq.size(),
-                           "",
-                           name));
+    add_annotation(name, "", *seq_start_i, *seq_start_i+a_seq.size()); 
   }
 }
 
@@ -1067,8 +1024,10 @@ void Sequence::find_sequences(std::list<Sequence>::iterator start,
 
 std::ostream& operator<<(std::ostream& out, const Sequence& s)
 {
-  for(Sequence::const_iterator s_i = s.begin(); s_i != s.end(); ++s_i) {
-    out << *s_i;
+  if (s.seq) {
+    for(Sequence::const_iterator s_i = s.begin(); s_i != s.end(); ++s_i) {
+      out << *s_i;
+    }
   }
   return out;
 }
@@ -1099,28 +1058,49 @@ bool operator<(const Sequence& x, const Sequence& y)
   }
 }
 
-bool operator==(const Sequence& x, const Sequence& y) 
+template <typename Iter1, typename Iter2>
+static
+bool sequence_insensitive_equality(Iter1 abegin, Iter1 aend, Iter2 bbegin, Iter2 bend)
 {
-  if (x.empty() and y.empty()) {
-    // if there's no sequence in either sequence structure, they're equal
+  Iter1 aseq_i = abegin;
+  Iter2 bseq_i = bbegin;
+  if (aend-abegin == bend-bbegin) {
+    // since the length of the two sequences is equal, we only need to
+    // test one.
+    for(; aseq_i != aend; ++aseq_i, ++bseq_i) {
+      if (toupper(*aseq_i) != toupper(*bseq_i)) {
+        return false;
+      }
+    }
     return true;
-  } else if (x.empty() or y.empty()) {
-    // if we fail the first test, and we discover one is empty,
-    // we know they can't be equal. (and we need to do this
-    // to prevent dereferencing an empty pointer)
-    return false;
-  } else if (x.seq_count != y.seq_count) {
-    // if they're of different lenghts, they're not equal
+  } else {
     return false;
   }
-  Sequence::const_iterator xseq_i = x.begin();
-  Sequence::const_iterator yseq_i = y.begin();
-  // since the length of the two sequences is equal, we only need to
-  // test one.
-  for(; xseq_i != x.end(); ++xseq_i, ++yseq_i) {
-    if (*xseq_i != *yseq_i) {
-      return false;
+}
+
+bool operator==(const Sequence& x, const Sequence& y) 
+{
+  if (x.seq and y.seq) {
+    // both x and y are defined
+    if (SeqSpan::isFamily(x.seq, y.seq)) {
+      // both are part of the same SeqSpan tree
+      return *(x.seq) == *(y.seq);
+    } else {
+      // we'll have to do a real comparison
+      return sequence_insensitive_equality<SeqSpan::const_iterator, SeqSpan::const_iterator>(
+               x.begin(), x.end(),
+               y.begin(), y.end()
+             );
     }
+  } else {
+    // true if they're both empty (with either a null SeqSpanRef or 
+    // a zero length string
+    return (x.size() == y.size());
   }
-  return true;
 }
+
+bool operator!=(const Sequence& x, const Sequence& y) 
+{
+  return not operator==(x, y);
+}
+