]> git.donarmstrong.com Git - mothur.git/blobdiff - trimseqscommand.cpp
sens.spec changes
[mothur.git] / trimseqscommand.cpp
index ef7dfdd0f45d649cc6296d91e106e75b9a6fd167..cbf5e13603ced805d185196fad9129eacb573d22 100644 (file)
@@ -8,6 +8,7 @@
  */
 
 #include "trimseqscommand.h"
+#include "needlemanoverlap.hpp"
 
 //***************************************************************************************************************
 
@@ -15,6 +16,7 @@ TrimSeqsCommand::TrimSeqsCommand(string option)  {
        try {
                
                abort = false;
+               comboStarts = 0;
                
                //allow user to run help
                if(option == "help") { help(); abort = true; }
@@ -22,7 +24,7 @@ TrimSeqsCommand::TrimSeqsCommand(string option)  {
                else {
                        //valid paramters for this command
                        string AlignArray[] =  {"fasta", "flip", "oligos", "maxambig", "maxhomop", "minlength", "maxlength", "qfile", 
-                                                                       "qthreshold", "qaverage", "allfiles", "qtrim", "processors", "outputdir","inputdir"};
+                                                                       "qthreshold", "qaverage", "allfiles", "qtrim","tdiffs", "pdiffs", "bdiffs", "processors", "outputdir","inputdir"};
                        
                        vector<string> myArray (AlignArray, AlignArray+(sizeof(AlignArray)/sizeof(string)));
                        
@@ -103,6 +105,18 @@ TrimSeqsCommand::TrimSeqsCommand(string option)  {
                        temp = validParameter.validFile(parameters, "maxlength", false);        if (temp == "not found") { temp = "0"; }
                        convert(temp, maxLength);
                        
+                       
+                       temp = validParameter.validFile(parameters, "bdiffs", false);           if (temp == "not found") { temp = "0"; }
+                       convert(temp, bdiffs);
+                       
+                       temp = validParameter.validFile(parameters, "pdiffs", false);           if (temp == "not found") { temp = "0"; }
+                       convert(temp, pdiffs);
+                       
+                       temp = validParameter.validFile(parameters, "tdiffs", false);           if (temp == "not found") { int tempTotal = pdiffs + bdiffs;  temp = toString(tempTotal); }
+                       convert(temp, tdiffs);
+                       
+                       if(tdiffs == 0){        tdiffs = bdiffs + pdiffs;       }
+                       
                        temp = validParameter.validFile(parameters, "qfile", true);     
                        if (temp == "not found")        {       qFileName = "";         }
                        else if(temp == "not open")     {       abort = true;           }
@@ -148,14 +162,17 @@ TrimSeqsCommand::TrimSeqsCommand(string option)  {
 void TrimSeqsCommand::help(){
        try {
                m->mothurOut("The trim.seqs command reads a fastaFile and creates .....\n");
-               m->mothurOut("The trim.seqs command parameters are fasta, flip, oligos, maxambig, maxhomop, minlength, maxlength, qfile, qthreshold, qaverage, qtrim and allfiles.\n");
+               m->mothurOut("The trim.seqs command parameters are fasta, flip, oligos, maxambig, maxhomop, minlength, maxlength, qfile, qthreshold, qaverage, diffs, qtrim and allfiles.\n");
                m->mothurOut("The fasta parameter is required.\n");
-               m->mothurOut("The flip parameter .... The default is 0.\n");
+               m->mothurOut("The flip parameter will output the reverse compliment of your trimmed sequence. The default is false.\n");
                m->mothurOut("The oligos parameter .... The default is "".\n");
                m->mothurOut("The maxambig parameter .... The default is -1.\n");
                m->mothurOut("The maxhomop parameter .... The default is 0.\n");
                m->mothurOut("The minlength parameter .... The default is 0.\n");
                m->mothurOut("The maxlength parameter .... The default is 0.\n");
+               m->mothurOut("The tdiffs parameter is used to specify the total number of differences allowed in the sequence. The default is pdiffs + bdiffs.\n");
+               m->mothurOut("The bdiffs parameter is used to specify the number of differences allowed in the barcode. The default is 0.\n");
+               m->mothurOut("The pdiffs parameter is used to specify the number of differences allowed in the primer. The default is 0.\n");
                m->mothurOut("The qfile parameter .....\n");
                m->mothurOut("The qthreshold parameter .... The default is 0.\n");
                m->mothurOut("The qaverage parameter .... The default is 0.\n");
@@ -208,8 +225,9 @@ int TrimSeqsCommand::execute(){
                #if defined (__APPLE__) || (__MACH__) || (linux) || (__linux)
                                if(processors == 1){
                                        ifstream inFASTA;
+                                       int numSeqs;
                                        openInputFile(fastaFile, inFASTA);
-                                       int numSeqs=count(istreambuf_iterator<char>(inFASTA),istreambuf_iterator<char>(), '>');
+                                       getNumSeqs(inFASTA, numSeqs);
                                        inFASTA.close();
                                        
                                        lines.push_back(new linePair(0, numSeqs));
@@ -250,35 +268,51 @@ int TrimSeqsCommand::execute(){
                                if (m->control_pressed) {  return 0; }
                #else
                                ifstream inFASTA;
-                               openInputFile(fastafileNames[s], inFASTA);
-                               numSeqs=count(istreambuf_iterator<char>(inFASTA),istreambuf_iterator<char>(), '>');
+                               int numSeqs;
+                               openInputFile(fastaFile, inFASTA);
+                               getNumSeqs(inFASTA, numSeqs);
                                inFASTA.close();
                                
                                lines.push_back(new linePair(0, numSeqs));
                                
-                               driverCreateSummary(fastafile, qFileName, trimSeqFile, scrapSeqFile, groupFile, fastaFileNames, lines[0], lines[0]);
+                               driverCreateTrim(fastaFile, qFileName, trimSeqFile, scrapSeqFile, groupFile, fastaFileNames, lines[0], lines[0]);
                                
                                if (m->control_pressed) {  return 0; }
                #endif
                                                
                                                                                
                for(int i=0;i<fastaFileNames.size();i++){
-                       ifstream inFASTA;
-                       string seqName;
-                       openInputFile(getRootName(fastaFile) + groupVector[i] + ".fasta", inFASTA);
-                       ofstream outGroups;
-                       openOutputFile(outputDir + getRootName(getSimpleName(fastaFile)) + groupVector[i] + ".groups", outGroups);
-                       outputNames.push_back(outputDir + getRootName(getSimpleName(fastaFile)) + groupVector[i] + ".groups");
-                       
-                       while(!inFASTA.eof()){
-                               if(inFASTA.get() == '>'){
-                                       inFASTA >> seqName;
-                                       outGroups << seqName << '\t' << groupVector[i] << endl;
+                       if (isBlank(fastaFileNames[i])) { remove(fastaFileNames[i].c_str()); }
+                       else if (filesToRemove.count(fastaFileNames[i]) > 0) { remove(fastaFileNames[i].c_str()); }
+                       else {
+                               ifstream inFASTA;
+                               string seqName;
+                               //openInputFile(getRootName(fastaFile) +  groupVector[i] + ".fasta", inFASTA);
+                               openInputFile(fastaFileNames[i], inFASTA);
+                               ofstream outGroups;
+                               string outGroupFilename = outputDir + getRootName(getSimpleName(fastaFileNames[i])) + "groups";
+                               openOutputFile(outGroupFilename, outGroups);
+                               //openOutputFile(outputDir + getRootName(getSimpleName(fastaFile)) + groupVector[i] + ".groups", outGroups);
+                               outputNames.push_back(outGroupFilename);
+                               
+                               string thisGroup = "";
+                               if (i > comboStarts) {
+                                       map<string, int>::iterator itCombo;
+                                       for(itCombo=combos.begin();itCombo!=combos.end(); itCombo++){
+                                               if(itCombo->second == i){       thisGroup = itCombo->first;     combos.erase(itCombo);  break;  }
+                                       }
+                               }else{ thisGroup = groupVector[i]; }
+                               
+                               while(!inFASTA.eof()){
+                                       if(inFASTA.get() == '>'){
+                                               inFASTA >> seqName;
+                                               outGroups << seqName << '\t' << thisGroup << endl;
+                                       }
+                                       while (!inFASTA.eof())  {       char c = inFASTA.get(); if (c == 10 || c == 13){        break;  }       }
                                }
-                               while (!inFASTA.eof())  {       char c = inFASTA.get(); if (c == 10 || c == 13){        break;  }       }
+                               outGroups.close();
+                               inFASTA.close();
                        }
-                       outGroups.close();
-                       inFASTA.close();
                }
                
                if (m->control_pressed) { 
@@ -315,7 +349,11 @@ int TrimSeqsCommand::driverCreateTrim(string filename, string qFileName, string
                if (oligoFile != "") {          
                        openOutputFile(groupFile, outGroups);   
                        for (int i = 0; i < fastaNames.size(); i++) {
+                       #if defined (__APPLE__) || (__MACH__) || (linux) || (__linux)
                                fastaFileNames.push_back(new ofstream((fastaNames[i] + toString(getpid()) + ".temp").c_str(), ios::ate)); 
+                       #else
+                               fastaFileNames.push_back(new ofstream((fastaNames[i] + toString(i) + ".temp").c_str(), ios::ate));                      
+                       #endif
                        }
                }
                
@@ -331,47 +369,47 @@ int TrimSeqsCommand::driverCreateTrim(string filename, string qFileName, string
                for(int i=0;i<line->num;i++){
                                
                        if (m->control_pressed) { 
-                               inFASTA.close(); 
-                               outFASTA.close();
-                               scrapFASTA.close();
-                               if (oligoFile != "") {   outGroups.close();   }
-                               if(qFileName != "")     {       qFile.close();  }
-                               for(int i=0;i<fastaFileNames.size();i++){
-                                       fastaFileNames[i]->close();
-                                       delete fastaFileNames[i];
-                               }       
+                               inFASTA.close(); outFASTA.close(); scrapFASTA.close(); if (oligoFile != "") {    outGroups.close();   } if(qFileName != "")     {       qFile.close();  }
+                               for(int i=0;i<fastaFileNames.size();i++){  fastaFileNames[i]->close(); delete fastaFileNames[i];  }     
                                for (int i = 0; i < outputNames.size(); i++) {  remove(outputNames[i].c_str()); }
                                return 0;
                        }
                        
-                       bool success = 1;
+                       int success = 1;
                        
                        Sequence currSeq(inFASTA);
 
                        string origSeq = currSeq.getUnaligned();
                        if (origSeq != "") {
-                               int group;
+                               int groupBar, groupPrime;
                                string trashCode = "";
+                               int currentSeqsDiffs = 0;
                                
                                if(qFileName != ""){
                                        if(qThreshold != 0)             {       success = stripQualThreshold(currSeq, qFile);   }
                                        else if(qAverage != 0)  {       success = cullQualAverage(currSeq, qFile);              }
-                                       if ((!qtrim) && (origSeq.length() != currSeq.getUnaligned().length())) { 
+                                       
+                                       if (qtrim == 1 && (origSeq.length() != currSeq.getUnaligned().length())) { 
                                                success = 0; //if you don't want to trim and the sequence does not meet quality requirements, move to scrap
                                        }
+
                                        if(!success)                    {       trashCode += 'q';                                                               }
                                }
                        
                                if(barcodes.size() != 0){
-                                       success = stripBarcode(currSeq, group);
-                                       if(!success){   trashCode += 'b';       }
+                                       success = stripBarcode(currSeq, groupBar);
+                                       if(success > bdiffs){   trashCode += 'b';       }
+                                       else{ currentSeqsDiffs += success;  }
                                }
-                       
+
                                if(numFPrimers != 0){
-                                       success = stripForward(currSeq);
-                                       if(!success){   trashCode += 'f';       }
+                                       success = stripForward(currSeq, groupPrime);
+                                       if(success > pdiffs){   trashCode += 'f';       }
+                                       else{ currentSeqsDiffs += success;  }
                                }
-                                       
+                               
+                               if (currentSeqsDiffs > tdiffs) { trashCode += 't';   }
+
                                if(numRPrimers != 0){
                                        success = stripReverse(currSeq);
                                        if(!success){   trashCode += 'r';       }
@@ -393,19 +431,28 @@ int TrimSeqsCommand::driverCreateTrim(string filename, string qFileName, string
                                if(flip){       currSeq.reverseComplement();    }               // should go last                       
                                
                                if(trashCode.length() == 0){
-                                       currSeq.setAligned(currSeq.getUnaligned());  //this is because of a modification we made to the sequence class to fix a bug.  all seqs have an aligned version, which is the version that gets printed.
+                                       currSeq.setAligned(currSeq.getUnaligned());
                                        currSeq.printSequence(outFASTA);
                                        if(barcodes.size() != 0){
-                                               outGroups << currSeq.getName() << '\t' << groupVector[group] << endl;
-                                               
+                                               string thisGroup = groupVector[groupBar];
+                                               int indexToFastaFile = groupBar;
+                                               if (primers.size() != 0){
+                                                       //does this primer have a group
+                                                       if (groupVector[groupPrime] != "") {  
+                                                               thisGroup += "." + groupVector[groupPrime]; 
+                                                               indexToFastaFile = combos[thisGroup];
+                                                       }
+                                               }
+                                               outGroups << currSeq.getName() << '\t' << thisGroup << endl;
                                                if(allFiles){
-                                                       currSeq.printSequence(*fastaFileNames[group]);                                  
+                                                       currSeq.printSequence(*fastaFileNames[indexToFastaFile]);                                       
                                                }
                                        }
                                }
                                else{
                                        currSeq.setName(currSeq.getName() + '|' + trashCode);
                                        currSeq.setUnaligned(origSeq);
+                                       currSeq.setAligned(origSeq);
                                        currSeq.printSequence(scrapFASTA);
                                }
                        }
@@ -532,14 +579,18 @@ void TrimSeqsCommand::getOligos(vector<string>& outFASTAVec){ //vector<ofstream*
                
                string type, oligo, group;
                int index=0;
+               //int indexPrimer = 0;
                
                while(!inOligos.eof()){
                        inOligos >> type;
-                       
+                                       
                        if(type[0] == '#'){
                                while (!inOligos.eof()) {       char c = inOligos.get(); if (c == 10 || c == 13){       break;  }       } // get rest of line if there's any crap there
                        }
                        else{
+                               //make type case insensitive
+                               for(int i=0;i<type.length();i++){       type[i] = toupper(type[i]);  }
+                               
                                inOligos >> oligo;
                                
                                for(int i=0;i<oligo.length();i++){
@@ -547,31 +598,75 @@ void TrimSeqsCommand::getOligos(vector<string>& outFASTAVec){ //vector<ofstream*
                                        if(oligo[i] == 'U')     {       oligo[i] = 'T'; }
                                }
                                
-                               if(type == "forward"){
-                                       forPrimer.push_back(oligo);
+                               if(type == "FORWARD"){
+                                       group = "";
+                                       
+                                       // get rest of line in case there is a primer name
+                                       while (!inOligos.eof()) {       
+                                               char c = inOligos.get(); 
+                                               if (c == 10 || c == 13){        break;  }
+                                               else if (c == 32 || c == 9){;} //space or tab
+                                               else {  group += c;  }
+                                       } 
+                                       
+                                       //check for repeat barcodes
+                                       map<string, int>::iterator itPrime = primers.find(oligo);
+                                       if (itPrime != primers.end()) { m->mothurOut("primer " + oligo + " is in your oligos file already."); m->mothurOutEndLine();  }
+                                       
+                                       primers[oligo]=index; index++;
+                                       groupVector.push_back(group);
+                                       
+                                       if(allFiles){
+                                               outFASTAVec.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(index) + "." + group + ".fasta"));
+                                               if (group == "") { //if there is not a group for this primer, then this file will not get written to, but we add it to keep the indexes correct
+                                                       filesToRemove.insert((outputDir + getRootName(getSimpleName(fastaFile)) + toString(index) + "." + group + ".fasta"));
+                                               }else {
+                                                       outputNames.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(index) + "." + group + ".fasta"));
+                                               }
+                                       }
+
                                }
-                               else if(type == "reverse"){
+                               else if(type == "REVERSE"){
                                        Sequence oligoRC("reverse", oligo);
                                        oligoRC.reverseComplement();
                                        revPrimer.push_back(oligoRC.getUnaligned());
                                }
-                               else if(type == "barcode"){
+                               else if(type == "BARCODE"){
                                        inOligos >> group;
-                                       barcodes[oligo]=index++;
+                                       
+                                       //check for repeat barcodes
+                                       map<string, int>::iterator itBar = barcodes.find(oligo);
+                                       if (itBar != barcodes.end()) { m->mothurOut("barcode " + oligo + " is in your oligos file already."); m->mothurOutEndLine();  }
+                                       
+                                       barcodes[oligo]=index; index++;
                                        groupVector.push_back(group);
                                        
                                        if(allFiles){
-                                               //outFASTAVec.push_back(new ofstream((outputDir + getRootName(getSimpleName(fastaFile)) + group + ".fasta").c_str(), ios::ate));
-                                               outputNames.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + group + ".fasta"));
-                                               outFASTAVec.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + group + ".fasta"));
+                                               outputNames.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(index) + "." + group + ".fasta"));
+                                               outFASTAVec.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(index) + "." + group + ".fasta"));
                                        }
-                               }
+                               }else{  m->mothurOut(type + " is not recognized as a valid type. Choices are forward, reverse, and barcode. Ignoring " + oligo + "."); m->mothurOutEndLine();  }
                        }
+                       gobble(inOligos);
                }
                
                inOligos.close();
                
-               numFPrimers = forPrimer.size();
+               //add in potential combos
+               if(allFiles){
+                       comboStarts = outFASTAVec.size()-1;
+                       for (map<string, int>::iterator itBar = barcodes.begin(); itBar != barcodes.end(); itBar++) {
+                               for (map<string, int>::iterator itPrime = primers.begin(); itPrime != primers.end(); itPrime++) {
+                                       if (groupVector[itPrime->second] != "") { //there is a group for this primer
+                                               outputNames.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(itBar->second) + "." + groupVector[itBar->second] + "." + toString(itPrime->second) + "." + groupVector[itPrime->second] + ".fasta"));
+                                               outFASTAVec.push_back((outputDir + getRootName(getSimpleName(fastaFile)) + toString(itBar->second) + "." + groupVector[itBar->second] + "." + toString(itPrime->second) + "." + groupVector[itPrime->second] + ".fasta"));
+                                               combos[(groupVector[itBar->second] + "." + groupVector[itPrime->second])] = outFASTAVec.size()-1;
+                                       }
+                               }
+                       }
+               }
+               
+               numFPrimers = primers.size();
                numRPrimers = revPrimer.size();
                
        }
@@ -579,30 +674,116 @@ void TrimSeqsCommand::getOligos(vector<string>& outFASTAVec){ //vector<ofstream*
                m->errorOut(e, "TrimSeqsCommand", "getOligos");
                exit(1);
        }
-
 }
-
 //***************************************************************************************************************
 
-bool TrimSeqsCommand::stripBarcode(Sequence& seq, int& group){
+int TrimSeqsCommand::stripBarcode(Sequence& seq, int& group){
        try {
+               
                string rawSequence = seq.getUnaligned();
-               bool success = 0;       //guilty until proven innocent
+               int success = bdiffs + 1;       //guilty until proven innocent
                
+               //can you find the barcode
                for(map<string,int>::iterator it=barcodes.begin();it!=barcodes.end();it++){
                        string oligo = it->first;
                        if(rawSequence.length() < oligo.length()){      //let's just assume that the barcodes are the same length
-                               success = 0;
-                               break;
+                               success = bdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
                        }
                        
                        if(compareDNASeq(oligo, rawSequence.substr(0,oligo.length()))){
                                group = it->second;
                                seq.setUnaligned(rawSequence.substr(oligo.length()));
-                               success = 1;
+                               success = 0;
                                break;
                        }
                }
+               
+               //if you found the barcode or if you don't want to allow for diffs
+//             cout << success;
+               if ((bdiffs == 0) || (success == 0)) { return success;  }
+               
+               else { //try aligning and see if you can find it
+//                     cout << endl;
+
+                       int maxLength = 0;
+
+                       Alignment* alignment;
+                       if (barcodes.size() > 0) {
+                               map<string,int>::iterator it=barcodes.begin();
+
+                               for(it;it!=barcodes.end();it++){
+                                       if(it->first.length() > maxLength){
+                                               maxLength = it->first.length();
+                                       }
+                               }
+                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+bdiffs+1));  
+
+                       }else{ alignment = NULL; } 
+                       
+                       //can you find the barcode
+                       int minDiff = 1e6;
+                       int minCount = 1;
+                       int minGroup = -1;
+                       int minPos = 0;
+                       
+                       for(map<string,int>::iterator it=barcodes.begin();it!=barcodes.end();it++){
+                               string oligo = it->first;
+//                             int length = oligo.length();
+                               
+                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                                       success = bdiffs + 10;
+                                       break;
+                               }
+                               
+                               //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                               alignment->align(oligo, rawSequence.substr(0,oligo.length()+bdiffs));
+                               oligo = alignment->getSeqAAln();
+                               string temp = alignment->getSeqBAln();
+               
+                               int alnLength = oligo.length();
+                               
+                               for(int i=oligo.length()-1;i>=0;i--){
+                                       if(oligo[i] != '-'){    alnLength = i+1;        break;  }
+                               }
+                               oligo = oligo.substr(0,alnLength);
+                               temp = temp.substr(0,alnLength);
+                               
+                               int newStart=0;
+                               int numDiff = countDiffs(oligo, temp);
+                               
+//                             cout << oligo << '\t' << temp << '\t' << numDiff << endl;                               
+                               
+                               if(numDiff < minDiff){
+                                       minDiff = numDiff;
+                                       minCount = 1;
+                                       minGroup = it->second;
+                                       minPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       minPos++;
+                                               }
+                                       }
+                               }
+                               else if(numDiff == minDiff){
+                                       minCount++;
+                               }
+
+                       }
+
+                       if(minDiff > bdiffs)    {       success = minDiff;              }       //no good matches
+                       else if(minCount > 1)   {       success = bdiffs + 100; }       //can't tell the difference between multiple barcodes
+                       else{                                                                                                   //use the best match
+                               group = minGroup;
+                               seq.setUnaligned(rawSequence.substr(minPos));
+                               success = minDiff;
+                       }
+                       
+                       if (alignment != NULL) {  delete alignment;  }
+                       
+               }
+//             cout << success << endl;
+               
                return success;
                
        }
@@ -615,28 +796,113 @@ bool TrimSeqsCommand::stripBarcode(Sequence& seq, int& group){
 
 //***************************************************************************************************************
 
-bool TrimSeqsCommand::stripForward(Sequence& seq){
+int TrimSeqsCommand::stripForward(Sequence& seq, int& group){
        try {
                string rawSequence = seq.getUnaligned();
-               bool success = 0;       //guilty until proven innocent
+               int success = pdiffs + 1;       //guilty until proven innocent
                
-               for(int i=0;i<numFPrimers;i++){
-                       string oligo = forPrimer[i];
-                       
-                       if(rawSequence.length() < oligo.length()){
-                               success = 0;
-                               break;
+               //can you find the primer
+               for(map<string,int>::iterator it=primers.begin();it!=primers.end();it++){
+                       string oligo = it->first;
+                       if(rawSequence.length() < oligo.length()){      //let's just assume that the primers are the same length
+                               success = pdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
                        }
                        
                        if(compareDNASeq(oligo, rawSequence.substr(0,oligo.length()))){
+                               group = it->second;
                                seq.setUnaligned(rawSequence.substr(oligo.length()));
-                               success = 1;
+                               success = 0;
                                break;
                        }
                }
+
+               //if you found the barcode or if you don't want to allow for diffs
+//             cout << success;
+               if ((pdiffs == 0) || (success == 0)) { return success;  }
                
-               return success;
+               else { //try aligning and see if you can find it
+//                     cout << endl;
+
+                       int maxLength = 0;
+
+                       Alignment* alignment;
+                       if (primers.size() > 0) {
+                               map<string,int>::iterator it=primers.begin();
+
+                               for(it;it!=primers.end();it++){
+                                       if(it->first.length() > maxLength){
+                                               maxLength = it->first.length();
+                                       }
+                               }
+                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+pdiffs+1));  
+
+                       }else{ alignment = NULL; } 
+                       
+                       //can you find the barcode
+                       int minDiff = 1e6;
+                       int minCount = 1;
+                       int minGroup = -1;
+                       int minPos = 0;
+                       
+                       for(map<string,int>::iterator it=primers.begin();it!=primers.end();it++){
+                               string oligo = it->first;
+//                             int length = oligo.length();
+                               
+                               if(rawSequence.length() < maxLength){   
+                                       success = pdiffs + 100;
+                                       break;
+                               }
+                               
+                               //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                               alignment->align(oligo, rawSequence.substr(0,oligo.length()+pdiffs));
+                               oligo = alignment->getSeqAAln();
+                               string temp = alignment->getSeqBAln();
                
+                               int alnLength = oligo.length();
+                               
+                               for(int i=oligo.length()-1;i>=0;i--){
+                                       if(oligo[i] != '-'){    alnLength = i+1;        break;  }
+                               }
+                               oligo = oligo.substr(0,alnLength);
+                               temp = temp.substr(0,alnLength);
+                               
+                               int newStart=0;
+                               int numDiff = countDiffs(oligo, temp);
+                               
+//                             cout << oligo << '\t' << temp << '\t' << numDiff << endl;                               
+                               
+                               if(numDiff < minDiff){
+                                       minDiff = numDiff;
+                                       minCount = 1;
+                                       minGroup = it->second;
+                                       minPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       minPos++;
+                                               }
+                                       }
+                               }
+                               else if(numDiff == minDiff){
+                                       minCount++;
+                               }
+
+                       }
+
+                       if(minDiff > pdiffs)    {       success = minDiff;              }       //no good matches
+                       else if(minCount > 1)   {       success = pdiffs + 10;  }       //can't tell the difference between multiple primers
+                       else{                                                                                                   //use the best match
+                               group = minGroup;
+                               seq.setUnaligned(rawSequence.substr(minPos));
+                               success = minDiff;
+                       }
+                       
+                       if (alignment != NULL) {  delete alignment;  }
+                       
+               }
+               
+               return success;
+
        }
        catch(exception& e) {
                m->errorOut(e, "TrimSeqsCommand", "stripForward");
@@ -744,7 +1010,7 @@ bool TrimSeqsCommand::compareDNASeq(string oligo, string seq){
                for(int i=0;i<length;i++){
                        
                        if(oligo[i] != seq[i]){
-                               if(oligo[i] == 'A' || oligo[i] == 'T' || oligo[i] == 'G' || oligo[i] == 'C')    {       success = 0;    }
+                               if(oligo[i] == 'A' || oligo[i] == 'T' || oligo[i] == 'G' || oligo[i] == 'C')    {       success = 0;    }
                                else if((oligo[i] == 'N' || oligo[i] == 'I') && (seq[i] == 'N'))                                {       success = 0;    }
                                else if(oligo[i] == 'R' && (seq[i] != 'A' && seq[i] != 'G'))                                    {       success = 0;    }
                                else if(oligo[i] == 'Y' && (seq[i] != 'C' && seq[i] != 'T'))                                    {       success = 0;    }
@@ -757,7 +1023,7 @@ bool TrimSeqsCommand::compareDNASeq(string oligo, string seq){
                                else if(oligo[i] == 'H' && (seq[i] != 'A' && seq[i] != 'T' && seq[i] != 'C'))   {       success = 0;    }
                                else if(oligo[i] == 'V' && (seq[i] != 'A' && seq[i] != 'C' && seq[i] != 'G'))   {       success = 0;    }                       
                                
-                               if(success == 0)        {       break;  }
+                               if(success == 0)        {       break;   }
                        }
                        else{
                                success = 1;
@@ -772,17 +1038,80 @@ bool TrimSeqsCommand::compareDNASeq(string oligo, string seq){
        }
 
 }
+//***************************************************************************************************************
 
+int TrimSeqsCommand::countDiffs(string oligo, string seq){
+       try {
+
+               int length = oligo.length();
+               int countDiffs = 0;
+               
+               for(int i=0;i<length;i++){
+                                                               
+                       if(oligo[i] != seq[i]){
+                               if(oligo[i] == 'A' || oligo[i] == 'T' || oligo[i] == 'G' || oligo[i] == 'C' || oligo[i] == '-' || oligo[i] == '.')      {       countDiffs++;   }
+                               else if((oligo[i] == 'N' || oligo[i] == 'I') && (seq[i] == 'N'))                                {       countDiffs++;   }
+                               else if(oligo[i] == 'R' && (seq[i] != 'A' && seq[i] != 'G'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'Y' && (seq[i] != 'C' && seq[i] != 'T'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'M' && (seq[i] != 'C' && seq[i] != 'A'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'K' && (seq[i] != 'T' && seq[i] != 'G'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'W' && (seq[i] != 'T' && seq[i] != 'A'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'S' && (seq[i] != 'C' && seq[i] != 'G'))                                    {       countDiffs++;   }
+                               else if(oligo[i] == 'B' && (seq[i] != 'C' && seq[i] != 'T' && seq[i] != 'G'))   {       countDiffs++;   }
+                               else if(oligo[i] == 'D' && (seq[i] != 'A' && seq[i] != 'T' && seq[i] != 'G'))   {       countDiffs++;   }
+                               else if(oligo[i] == 'H' && (seq[i] != 'A' && seq[i] != 'T' && seq[i] != 'C'))   {       countDiffs++;   }
+                               else if(oligo[i] == 'V' && (seq[i] != 'A' && seq[i] != 'C' && seq[i] != 'G'))   {       countDiffs++;   }                       
+                               
+                       }
+               }
+               
+               return countDiffs;
+       }
+       catch(exception& e) {
+               m->errorOut(e, "TrimSeqsCommand", "countDiffs");
+               exit(1);
+       }
+
+}
 //***************************************************************************************************************
 
 bool TrimSeqsCommand::stripQualThreshold(Sequence& seq, ifstream& qFile){
        try {
+//             string rawSequence = seq.getUnaligned();
+//             int seqLength;  // = rawSequence.length();
+//             string name, temp, temp2;
+//             
+//             qFile >> name;
+//             
+//             //get rest of line
+//             temp = "";
+//             while (!qFile.eof())    {       
+//                     char c = qFile.get(); 
+//                     if (c == 10 || c == 13){        break;  }       
+//                     else { temp += c; }
+//             } 
+//     
+//             int pos = temp.find("length");
+//             if (pos == temp.npos) { m->mothurOut("Cannot find length in qfile for " + seq.getName()); m->mothurOutEndLine();  seqLength = 0;  }
+//             else {
+//                     string tempLength = temp.substr(pos);
+//                     istringstream iss (tempLength,istringstream::in);
+//                     iss >> temp;
+//             }
+//             
+//             splitAtEquals(temp2, temp); //separates length=242, temp=length, temp2=242
+//             convert(temp, seqLength); //converts string to int
+//     
+//             if (name.length() != 0) {  if(name.substr(1) != seq.getName())  {       m->mothurOut("sequence name mismatch btwn fasta and qual file"); m->mothurOutEndLine(); }  } 
+               
                string rawSequence = seq.getUnaligned();
-               int seqLength = rawSequence.length();
+               int seqLength = seq.getNumBases();
+               bool success = 0;       //guilty until proven innocent
                string name;
                
                qFile >> name;
-               if (name.length() != 0) {  if(name.substr(1) != seq.getName())  {       m->mothurOut("sequence name mismatch btwn fasta and qual file"); m->mothurOutEndLine(); }  } 
+               if (name[0] == '>') {  if(name.substr(1) != seq.getName())      {       m->mothurOut("sequence name mismatch btwn fasta: " + seq.getName() + " and qual file: " + name); m->mothurOutEndLine(); }  }
+               
                while (!qFile.eof())    {       char c = qFile.get(); if (c == 10 || c == 13){  break;  }       }
                
                int score;
@@ -791,7 +1120,7 @@ bool TrimSeqsCommand::stripQualThreshold(Sequence& seq, ifstream& qFile){
                for(int i=0;i<seqLength;i++){
                        qFile >> score;
                        
-                       if(score <= qThreshold){
+                       if(score < qThreshold){
                                end = i;
                                break;
                        }