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//Copyright inria / irisa (2013)
//
// this file is shared between MindTheGap and Kissnp2,
// and possibly other Minia-related software
//
//raluca.uricaru@gmail.com
//pierre.peterlongo@inria.fr
//guillaume.rizk@irisa.fr
//rayan.chikhi@ens-cachan.org
//
//This software is governed by the CeCILL license under French law and
//abiding by the rules of distribution of free software. You can use,
//modify and/ or redistribute the software under the terms of the CeCILL
//license as circulated by CEA, CNRS and INRIA at the following URL
//"http://www.cecill.info".
//
//As a counterpart to the access to the source code and rights to copy,
//modify and redistribute granted by the license, users are provided only
//with a limited warranty and the software's author, the holder of the
//economic rights, and the successive licensors have only limited
//liability.
//
//In this respect, the user's attention is drawn to the risks associated
//with loading, using, modifying and/or developing or reproducing the
//software by the user in light of its specific status of free software,
//that may mean that it is complicated to manipulate, and that also
//therefore means that it is reserved for developers and experienced
//professionals having in-depth computer knowledge. Users are therefore
//encouraged to load and test the software's suitability as regards their
//requirements in conditions enabling the security of their systems and/or
//data to be ensured and, more generally, to use and operate it in the
//same conditions as regards security.
//
//The fact that you are presently reading this means that you have had
//knowledge of the CeCILL license and that you accept its terms.
#include "IterativeExtensions.h"
//#define DONTMARK
AssocPairedSet *pairedBranchingKmers;
// this type bears similarity with Traversal.h:kmer_strand_nt but I wanted to rename "nt" to "depth" for clarity
struct kmer_strand_depth {
kmer_type kmer;
int strand;
int depth;
kmer_strand_depth(kmer_type kmer, int strand, int depth) : kmer(kmer), strand(strand), depth(depth) {}
bool operator<(const kmer_strand_depth &other) const { // needed for comparisons inside a list
if (kmer != other.kmer)
return (kmer < other.kmer);
if (depth != other.depth)
return depth < other.depth;
return (strand < other.strand);
}
};
/*
* our assembly graph is connected by (k-1)-overlaps,
* so this function is used to make sure we see each (k-1)-overlap in at most one right extremity
*/
bool compare_and_mark_last_k_minus_one_mer(string node, set<kmer_type> &kmers_set)
{
kmer_type kmer_fw, kmer_rc;
sizeKmer--;
kmer_type kmer = extractKmerFromRead( (char *)node.c_str(), node.size() - sizeKmer, &kmer_fw, &kmer_rc, false);
int strand = (kmer == kmer_rc);
// char kmer_seq[100];
// code2seq(kmer_fw,kmer_seq); // convert starting kmer to nucleotide seq
// //printf("checking marking kmer: %s\n",kmer_seq);
sizeKmer++;
if (kmers_set.find(kmer) != kmers_set.end())
return true;
kmers_set.insert(kmer);
return false;
}
// default extension modes (do not change)
IterativeExtensions::Traversal_type IterativeExtensions::traversal_type = Monument;
IterativeExtensions::When_to_stop_extending IterativeExtensions::when_to_stop_extending = Until_max_depth;
bool IterativeExtensions::dont_output_first_nucleotide = false;
void IterativeExtensions::construct_linear_seqs(string L, int max_depth, string output_file)
{
construct_linear_seqs(L, string(),max_depth,output_file,2,1000000000);
}
/*
* return the contig which starts with L, as well as all ths contigs that follow him, up to max_depth.
* results go to a fasta file (output_file)
*
* requires that:
* global variables terminator, bloo1 and false_positives are already constructed by minia
*
* outputs:
* a set of contigs in the output file */
void IterativeExtensions::construct_linear_seqs(string L,string R, int max_depth, string output_file , int verb, int max_nodes, parcours_t search_mode, bool swf )
{
bool debug = verb>=2 ? 1 : 0;
kmer_type kmer;
char kmer_seq[sizeKmer+1];
FILE *linear_seqs_file;
Traversal *traversal;
if (traversal_type == IterativeExtensions::SimplePaths)
traversal = new SimplePathsTraversal(bloo1,false_positives,terminator);
else if (traversal_type == IterativeExtensions::Monument)
traversal = new MonumentTraversal(bloo1,false_positives,terminator);
traversal->set_maxlen(max_depth);
traversal->set_max_depth(500);
traversal->set_max_breadth(20);
long long nbNodes = 0;
long long totalnt=0;
long long contig_len =0;
long long maxlen=1000000;
char *right_traversal = (char *) malloc(maxlen*sizeof(char));
char *node = (char *) malloc(maxlen*sizeof(char));
node [0] = '\0';
if(!output_file.empty())
{
linear_seqs_file = fopen((char * )output_file.c_str(),"w");
}
else {
linear_seqs_file = NULL;
IterativeExtensions::when_to_stop_extending = IterativeExtensions::After_first_contig; // sequence
}
STARTWALL(nodes);
// heuristics: start from the last kmer of L
vector < kmer_strand_depth > kmers_to_traverse;
kmer_type kmer_fw, kmer_rc;
kmer = extractKmerFromRead( (char *)L.c_str(), L.size() - sizeKmer, &kmer_fw, &kmer_rc, false);
int strand = (kmer == kmer_rc);
kmer_strand_depth first_kmer (kmer,strand,0);
kmers_to_traverse.push_back(first_kmer);
#ifndef DONTMARK
set<kmer_type> already_extended_from;
// compare_and_mark_last_k_minus_one_mer(L, already_extended_from); // mark first kmer to never extend from it again, // L will be marked at first iteration below
#endif
while (kmers_to_traverse.size() > 0) // min_depth is max_gap_length here
{
kmer_strand_depth ksd (0,0,0);
if (search_mode == PROFONDEUR)
{
ksd = kmers_to_traverse.back();
kmers_to_traverse.pop_back();
}
else if (search_mode == LARGEUR)
{
ksd = kmers_to_traverse.front();
kmers_to_traverse.erase (kmers_to_traverse.begin());
}
kmer = ksd.kmer;
int strand = ksd.strand;
int depth = ksd.depth;
code2seq(kmer,kmer_seq); // convert starting kmer to nucleotide seq
if (strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
if (debug)
printf(" --- iteration: kmer %s%s (of length %d) depth %d nbNodes explored %lli ---\n",kmer_seq,strand?" (internally rc)":"", (int)strlen(kmer_seq), depth,nbNodes);
// right extension
int len_right = traversal->traverse(kmer,right_traversal,strand);
if (debug)
printf("right traversal %i = %s\n", len_right,right_traversal);
// save the node
strcpy(node,kmer_seq);// + starting_kmer if we ask for the de bruijn graph or for a text output
strcat(node,right_traversal);// + right_traversal
int node_len=len_right+sizeKmer;
//TODO: watch the reason of wrong length with depth = 0
fprintf(linear_seqs_file,">%lli__len__%i__depth__%i\n",nbNodes,node_len,depth);
if (depth == 0 && dont_output_first_nucleotide )
/* we need this in mapsembler:
// the first used kmer should be a k-1 mer. Indeed, if a first kmer is extracted from a sequence :
/// -------------****** (k=6), then this node is the one linked to a new one starting with ******, thus with an overlap of k and not k-1. */
fprintf(linear_seqs_file,"%s\n",node+1);
else
fprintf(linear_seqs_file,"%s\n",node);
nbNodes++;
totalnt+=node_len;
if (debug)
{
// printf ("[L=%s] assembled a %d bp node at depth %d\n",L.c_str(),node_len, depth);
printf("(node seq: %s)\n",node);
}
// if we only want 1 extension, stop now
if (when_to_stop_extending == IterativeExtensions::After_first_contig)
{
if (debug)
printf("Stopping because we want only 1 extension\n");
break;
}
if(swf)
{
char * found = NULL;
found =strstr(node, R.c_str());
if(found!=NULL && depth > sizeKmer)
{
if (debug)
printf("swf STOP \n");
break;
}
}
if(nbNodes > max_nodes) //GR stop when too complex huum when to stop ?
{
if (debug)
printf("... XXX Stopped extending node %s because %lld nodes reached. Was at depth %d.\n", node, nbNodes, depth);
break;
}
// if max depth reached, don't extend that one
if (depth + node_len > max_depth)
{
if (debug)
printf("... XXX max depth reached for node %s (depth + node length %i %i = %i) \n",node, depth,node_len,depth + node_len );
continue;
}
#ifndef DONTMARK
// make sure this is the only time we see this (k-1)-overlap
bool already_seen = compare_and_mark_last_k_minus_one_mer(node, already_extended_from);
if (already_seen)
{
if (debug)
printf("... XXX not extending node %s becaues last k-1-mer was already seen\n",node);
continue;
}
#endif
kmer = extractKmerFromRead( node, node_len - sizeKmer, &kmer_fw, &kmer_rc, false);
strand = (kmer == kmer_rc);
if (debug)
{
code2seq(kmer,kmer_seq);
if (strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
printf("... restarting from kmer %s %s\n",kmer_seq,strand?"(internally rc)":"");
}
// continue extending from immediately overlapping kmers
// there may be just one 1 possibility (there was in-branching)
int nb_extensions = 0;
for(int test_nt=0; test_nt<4; test_nt++)
{
int current_strand = strand;
kmer_type current_kmer = next_kmer(kmer,test_nt, ¤t_strand);
if(bloo1->contains(current_kmer) && !false_positives->contains(current_kmer)){
kmers_to_traverse.push_back(kmer_strand_depth (current_kmer,current_strand,depth + len_right +1 )); // ou plutot depth + len_right +1 (+1 = la nt ajoutee ici) (et pas node_len) ?
nb_extensions++;
if (debug)
{
code2seq(current_kmer,kmer_seq); // convert starting kmer to nucleotide seq
if (current_strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
printf("... --> found kmer immediately after kmer %s: %s \n",kmer_seq,current_strand?"(internally rc)":"");
}
}
}
if (debug)
printf("... number of extensions: %d\n", nb_extensions);
}
//delete terminator;
//delete traversal;
free(right_traversal);
//SolidKmers->close();
free(node);
fclose(linear_seqs_file);
}
void IterativeExtensions::construct_linear_seqs_paired(string L, int max_depth, string output_file)
{
construct_linear_seqs_paired(L, string(),max_depth,output_file,2,1000000000);
}
/*
* return the contig which starts with L, as well as all ths contigs that follow him, up to max_depth.
* results go to a fasta file (output_file)
*
* requires that:
* global variables terminator, bloo1 and false_positives are already constructed by minia
*
* outputs:
* a set of contigs in the output file */
void IterativeExtensions::construct_linear_seqs_paired(string L,string R, int max_depth, string output_file , int verb, int max_nodes)
{
///code for paired kmer
int number_of_kmers_memorized = 1000; //should be greater than insert length, but maybe too much can be detrimental ?
kmer_type * kmer_history = (kmer_type *) malloc (sizeof(kmer_type)*number_of_kmers_memorized);
memset(kmer_history,0,sizeof(kmer_type)*number_of_kmers_memorized);
int store_index =0;
///
bool debug = verb>=2 ? 1 : 0;
char bin2NT[4] = {'A','C','T','G'};
char bin2NTrev[4] = {'T','G','A','C'};
char binrev[4] = {2,3,0,1};
kmer_type kmer;
char kmer_seq[sizeKmer+1];
FILE *linear_seqs_file;
Traversal *traversal;
if (traversal_type == IterativeExtensions::SimplePaths)
traversal = new SimplePathsTraversal(bloo1,false_positives,terminator);
else if (traversal_type == IterativeExtensions::Monument)
traversal = new MonumentTraversal(bloo1,false_positives,terminator);
traversal->set_maxlen(max_depth);
traversal->set_max_depth(500);
traversal->set_max_breadth(20);
long long nbNodes = 0;
long long totalnt=0;
long long contig_len =0;
long long maxlen=1000000;
char *right_traversal = (char *) malloc(maxlen*sizeof(char));
char *node = (char *) malloc(maxlen*sizeof(char));
node [0] = '\0';
if(!output_file.empty())
{
linear_seqs_file = fopen((char * )output_file.c_str(),"w");
}
else {
linear_seqs_file = NULL;
IterativeExtensions::when_to_stop_extending = IterativeExtensions::After_first_contig; // sequence
}
STARTWALL(nodes);
// heuristics: start from the last kmer of L
vector < kmer_strand_depth > kmers_to_traverse;
kmer_type kmer_fw, kmer_rc;
kmer = extractKmerFromRead( (char *)L.c_str(), L.size() - sizeKmer, &kmer_fw, &kmer_rc, false);
int strand = (kmer == kmer_rc);
kmer_strand_depth first_kmer (kmer,strand,0);
kmers_to_traverse.push_back(first_kmer);
#ifndef DONTMARK
set<kmer_type> already_extended_from;
compare_and_mark_last_k_minus_one_mer(L, already_extended_from); // mark first kmer to never extend from it again
#endif
while (kmers_to_traverse.size() > 0) // min_depth is max_gap_length here
{
kmer_strand_depth ksd = kmers_to_traverse.back();
kmers_to_traverse.pop_back();
kmer = ksd.kmer;
int strand = ksd.strand;
int depth = ksd.depth;
code2seq(kmer,kmer_seq); // convert starting kmer to nucleotide seq
if (strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
if (debug)
printf("iteration: kmer %s%s (of length %zu) depth %d nbNodes explored %lli\n",kmer_seq,strand?" (internally rc)":"", strlen(kmer_seq), depth,nbNodes);
// right extension
int len_right = traversal->traverse(kmer,right_traversal,strand);
if (debug)
printf("right traversal = %s\n", right_traversal);
// save the node
strcpy(node,kmer_seq);// + starting_kmer if we ask for the de bruijn graph or for a text output
strcat(node,right_traversal);// + right_traversal
int node_len=len_right+sizeKmer;
fprintf(linear_seqs_file,">%lli__len__%i__depth__%i\n",nbNodes,node_len,depth);
if (depth == 0 && dont_output_first_nucleotide )
/* we need this in mapsembler:
// the first used kmer should be a k-1 mer. Indeed, if a first kmer is extracted from a sequence :
/// -------------****** (k=6), then this node is the one linked to a new one starting with ******, thus with an overlap of k and not k-1. */
fprintf(linear_seqs_file,"%s\n",node+1);
else
fprintf(linear_seqs_file,"%s\n",node);
////memorize previous kmers in the kmer_history, it is a circular buffer
kmer_type kmer, graine, graine_revcomp ;
for (int ii=0; ii<node_len-sizeKmer+1; ii++)
{
kmer = extractKmerFromRead(right_traversal,ii,&graine,&graine_revcomp);
kmer_history[store_index]=kmer;
store_index = (store_index + 1) % number_of_kmers_memorized ;
}
nbNodes++;
totalnt+=node_len;
if (debug)
{
printf ("[L=%s] assembled a %d bp node at depth %d\n",L.c_str(),node_len, depth);
printf("(node seq: %s)\n",node);
}
// if we only want 1 extension, stop now
if (when_to_stop_extending == IterativeExtensions::After_first_contig)
break;
#ifdef STOPWHENFOUND
char * found = NULL;
found =strstr(node, R.c_str());
if(found!=NULL)
break;
#endif
if(nbNodes > max_nodes) //GR stop when too complex huum when to stop ?
break;
// if max depth reached, don't extend that one and clear kmer history, a new branch is explored
if (depth + node_len > max_depth)
{
memset(kmer_history,0,sizeof(kmer_type)*number_of_kmers_memorized);
store_index = 0;
continue;
}
#ifndef DONTMARK
// make sure this is the only time we see this (k-1)-overlap
bool already_seen = compare_and_mark_last_k_minus_one_mer(node, already_extended_from);
if (already_seen)
continue;
#endif
//this is the last kmer of the contig
kmer = extractKmerFromRead( node, node_len - sizeKmer, &kmer_fw, &kmer_rc, false);
strand = (kmer == kmer_rc);
if (debug)
{
code2seq(kmer,kmer_seq);
if (strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
printf("restarting from kmer %s %s\n",kmer_seq,strand?"(internally rc)":"");
}
// continue extending from immediately overlapping kmers
// there may be just one 1 possibility (there was in-branching)
//we are at the end of a contig, we push the starting point of all possible branches in a stack of kmers
//todo : push only kmers that are consistent with pairing information
//use kmer_history and the pairedBranchingKmers object for that
//Raluca
//first, compute the degree of the kmer named "kmer"
int degree = 0;
int current_strand;
kmer_type current_kmer, aux_kmer;
int nt1=-1, nt2=-1;
bool ok_nt1=false, ok_nt2=false;
for(int test_nt=0; test_nt<4; test_nt++)
{
current_strand = strand;
current_kmer = next_kmer(kmer,test_nt, ¤t_strand);
if(bloo1->contains(current_kmer) && !false_positives->contains(current_kmer))
{
degree++;
nt1==-1 ? nt1 = test_nt : nt2 = test_nt;
}
}
if(debug)
{
printf("degre %i \n",degree);
}
//second, depending on the degree and on whether we find the current_kmer in the history, we decide to push it or not
if ( degree == 0 ) // this path ends, a new one will be explored, clear kmer history
{
memset(kmer_history,0,sizeof(kmer_type)*number_of_kmers_memorized);
store_index = 0;
}
else
if ( degree == 1 ) // not branching, is this possible knowing that we're at the end of a contig?
{
//if it is possible, then simply push the kmer obtained with nt1
ok_nt1 = true;
ok_nt2 = false;
}
else
if (degree == 2 )
{
//we have the 2 extension, nt1 and nt2
//first, get the entries in pairedBranchingKmers for kmer named "kmer"
pair_nt_kmer_t val;
int is_present = pairedBranchingKmers->get(kmer, &val);
if (!is_present) // strange
{
//simply push the two possible branches
ok_nt1 = ok_nt2 = true;
}
else
{
if(debug)
{
printf("is_present, %c %c \n",bin2NT[nt1],bin2NT[nt2]);
}
int aux_nt;
//search each of the two nucleotides in val of kmer
for (int i=1; i<=2; i++)
{
aux_kmer = -1;
i==1 ? aux_nt = bin2NT[nt1] : aux_nt = bin2NT[nt2];
if(debug)
{
// char seq[100];
// code2seq(aux_kmer, seq);
printf("loop %i aux_nt %c \n",i,aux_nt);
}
if ( val.nk1.nt == aux_nt )
aux_kmer = val.nk1.prev_kmer;
else
if ( val.nk2.nt == aux_nt )
aux_kmer = val.nk2.prev_kmer;
else
{
i==1 ? ok_nt1 = false : ok_nt2 = false;
if ( i==2 && !ok_nt1 && !ok_nt2 )
break;
}
if(debug)
{
printf("loop %i ok_nt1 %i ok_nt2 %i \n",i,ok_nt1,ok_nt2);
}
//if one of the nt1 and nt2 corresponds, then search the corresponding prev_kmer (aux_kmer) in the history, if the prev_kmer is not present then ok_nt = false
if ( aux_kmer != -1 )
{
for ( int j=0; j<number_of_kmers_memorized; j++)
if ( kmer_history[j] == aux_kmer)
{
i==1 ? ok_nt1 = true : ok_nt2 = true;
if(debug)
{
char seq[100];
code2seq(aux_kmer, seq);
printf("found kmer ok_nt1 %i ok_nt2 %i %s \n",ok_nt1,ok_nt2,seq);
}
break;
}
// if both ok_nt false, then push the two of them
if ( i==2 && !ok_nt1 && !ok_nt2 )
ok_nt1 = ok_nt2 = true;
}
}
}
}
else //degree== 3 or 4
{
continue;
}
// if both ok_nt false, then push the two of them
if ( !ok_nt1 && !ok_nt2 )
ok_nt1 = ok_nt2 = true;
//else ???
// we cannot decide among the 3 or 4 knowing that our paired history keeps only 2 possible branchings per kmer TRUE? so we push them all?
if ( ok_nt1 )
{
current_strand = strand;
current_kmer = next_kmer(kmer, nt1, ¤t_strand);
kmers_to_traverse.push_back(kmer_strand_depth (current_kmer,current_strand,depth + node_len));
if (debug)
{
code2seq(current_kmer,kmer_seq); // convert starting kmer to nucleotide seq
if (current_strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
printf("push kmer immediately after: %s %s \n",kmer_seq,current_strand?"(internally rc)":"");
}
}
if ( ok_nt2 )
{
current_strand = strand;
current_kmer = next_kmer(kmer, nt2, ¤t_strand);
kmers_to_traverse.push_back(kmer_strand_depth (current_kmer,current_strand,depth + node_len));
if (debug)
{
code2seq(current_kmer,kmer_seq); // convert starting kmer to nucleotide seq
if (current_strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
printf("push kmer immediately after: %s %s \n",kmer_seq,current_strand?"(internally rc)":"");
}
}
}
free(kmer_history);
//delete terminator;
//delete traversal;
free(right_traversal);
//SolidKmers->close();
free(node);
fclose(linear_seqs_file);
}
/* requires:
* bloo1 and false_positives are already constructed by minia
*
* outputs:
* a pair containing:
* 1/ the extension
* 2/ the length of the first unitig composing this sequence.
* - if the traversal is a simple traversal, then this length is equal to the length of the extension
* - else (if the traversal is a monument traversal), then this length is equal to the starting position of the first bubble (if exist))
*/
pair<char *, int> IterativeExtensions::extend_a_snp(string L, int max_depth)
{
// printf("\nExtended sequ = %s\n", L.c_str()); // DEB
// bool debug = true;
bool debug = false;
kmer_type kmer;
char kmer_seq[sizeKmer+1];
Traversal *traversal;
if (traversal_type == IterativeExtensions::SimplePaths)
traversal = new SimplePathsTraversal(bloo1,false_positives,terminator);
else if (traversal_type == IterativeExtensions::Monument)
traversal = new MonumentTraversal(bloo1,false_positives,terminator);
traversal->set_maxlen(max_depth);
traversal->set_max_depth(500);
traversal->set_max_breadth(20);
long long nbNodes = 0;
long long totalnt=0;
long long contig_len =0;
long long maxlen=1000000;
char *right_traversal = (char *) malloc(maxlen*sizeof(char));
char *node = (char *) malloc(maxlen*sizeof(char));
node [0] = '\0';
IterativeExtensions::when_to_stop_extending = IterativeExtensions::After_first_contig; // sequence
STARTWALL(nodes);
// heuristics: start from the last kmer of L
kmer_type kmer_fw, kmer_rc;
kmer = extractKmerFromRead( (char *)L.c_str(), L.size() - sizeKmer, &kmer_fw, &kmer_rc, false);
int strand = (kmer == kmer_rc);
code2seq(kmer,kmer_seq); // convert starting kmer to nucleotide seq
// printf("Extended kmer = %s strand = %d\n", kmer_seq, strand); // DEB
if (strand == 1)
revcomp_sequence(kmer_seq,sizeKmer);
// if (debug)
// printf("iteration: kmer %s%s depth %d\n",kmer_seq,strand?" (internally rc)":"", 0);
// right extension
int len_right = traversal->traverse(kmer,right_traversal,strand);
// if(debug) printf("right traversal = %s\n", right_traversal);
// save the node
strcpy(node,kmer_seq);// + starting_kmer if we ask for the de bruijn graph or for a text output
strcat(node,right_traversal);// + right_traversal
free(right_traversal);
int first_divergence=strlen(node);
if (traversal_type == IterativeExtensions::Monument && traversal->bubbles_positions.size()>0) {
first_divergence=traversal->bubbles_positions[0].first+sizeKmer;
}
return std::make_pair(node,first_divergence);
}
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