]> git.donarmstrong.com Git - mothur.git/blobdiff - trimoligos.cpp
added order=A or order=B to trim.flows and shhh.flows commands. added load balancing...
[mothur.git] / trimoligos.cpp
index 2f92cc847ca75e1a0983aeef2d09e9eaab68e57f..a53f4e85388f0f7bbb6fda40c2074b4f555e29d3 100644 (file)
@@ -14,7 +14,7 @@
 
 /********************************************************************/
 //strip, pdiffs, bdiffs, primers, barcodes, revPrimers
-TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<string, int> br, map<string, int> rbr, vector<string> r, vector<string> lk, vector<string> sp){
+TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<string, int> br, vector<string> r, vector<string> lk, vector<string> sp){
        try {
                m = MothurOut::getInstance();
                
@@ -24,11 +24,37 @@ TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<stri
         sdiffs = s;
                
                barcodes = br;
-        rbarcodes = rbr;
                primers = pr;
                revPrimer = r;
         linker = lk;
         spacer = sp;
+        maxFBarcodeLength = 0;
+        for(map<string,int>::iterator it=barcodes.begin();it!=barcodes.end();it++){
+            if(it->first.length() > maxFBarcodeLength){
+                maxFBarcodeLength = it->first.length();
+            }
+        }
+        maxFPrimerLength = 0;
+        map<string,int>::iterator it; 
+        for(it=primers.begin();it!=primers.end();it++){
+            if(it->first.length() > maxFPrimerLength){
+                maxFPrimerLength = it->first.length();
+            }
+        }
+        
+        maxLinkerLength = 0;
+        for(int i = 0; i < linker.size(); i++){
+            if(linker[i].length() > maxLinkerLength){
+                maxLinkerLength = linker[i].length();
+            }
+        }
+        
+        maxSpacerLength = 0;
+        for(int i = 0; i < spacer.size(); i++){
+            if(spacer[i].length() > maxSpacerLength){
+                maxSpacerLength = spacer[i].length();
+            }
+        }
        }
        catch(exception& e) {
                m->errorOut(e, "TrimOligos", "TrimOligos");
@@ -37,7 +63,7 @@ TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<stri
 }
 /********************************************************************/
 //strip, pdiffs, bdiffs, primers, barcodes, revPrimers
-TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<string, int> br, vector<string> r, vector<string> lk, vector<string> sp){
+TrimOligos::TrimOligos(int p, int b, int l, int s, map<int, oligosPair> pr, map<int, oligosPair> br){
        try {
                m = MothurOut::getInstance();
                
@@ -46,11 +72,60 @@ TrimOligos::TrimOligos(int p, int b, int l, int s, map<string, int> pr, map<stri
         ldiffs = l;
         sdiffs = s;
                
-               barcodes = br;
-               primers = pr;
-               revPrimer = r;
-        linker = lk;
-        spacer = sp;
+        maxFBarcodeLength = 0;
+        for(map<int,oligosPair>::iterator it=br.begin();it!=br.end();it++){
+            string forward = it->second.forward;
+            map<string, vector<int> >::iterator itForward = ifbarcodes.find(forward);
+            
+            if(forward.length() > maxFBarcodeLength){ maxFBarcodeLength = forward.length();  }
+            
+            if (itForward == ifbarcodes.end()) {
+                vector<int> temp; temp.push_back(it->first);
+                ifbarcodes[forward] = temp;
+            }else { itForward->second.push_back(it->first); }
+        }
+
+        maxFPrimerLength = 0;
+        for(map<int,oligosPair>::iterator it=pr.begin();it!=pr.end();it++){
+            string forward = it->second.forward;
+            map<string, vector<int> >::iterator itForward = ifprimers.find(forward);
+            
+            if(forward.length() > maxFPrimerLength){ maxFPrimerLength = forward.length();  }
+            
+            if (itForward == ifprimers.end()) {
+                vector<int> temp; temp.push_back(it->first);
+                ifprimers[forward] = temp;
+            }else { itForward->second.push_back(it->first); }
+        }
+        
+        maxRBarcodeLength = 0;
+        for(map<int,oligosPair>::iterator it=br.begin();it!=br.end();it++){
+            string forward = it->second.reverse;
+            map<string, vector<int> >::iterator itForward = irbarcodes.find(forward);
+            
+            if(forward.length() > maxRBarcodeLength){ maxRBarcodeLength = forward.length();  }
+            
+            if (itForward == irbarcodes.end()) {
+                vector<int> temp; temp.push_back(it->first);
+                irbarcodes[forward] = temp;
+            }else { itForward->second.push_back(it->first); }
+        }
+        
+        maxRPrimerLength = 0;
+        for(map<int,oligosPair>::iterator it=pr.begin();it!=pr.end();it++){
+            string forward = it->second.reverse;
+            map<string, vector<int> >::iterator itForward = irprimers.find(forward);
+            
+            if(forward.length() > maxRPrimerLength){ maxRPrimerLength = forward.length();  }
+            
+            if (itForward == irprimers.end()) {
+                vector<int> temp; temp.push_back(it->first);
+                irprimers[forward] = temp;
+            }else { itForward->second.push_back(it->first); }
+        }
+
+        ipbarcodes = br;
+        ipprimers = pr;
        }
        catch(exception& e) {
                m->errorOut(e, "TrimOligos", "TrimOligos");
@@ -69,6 +144,24 @@ TrimOligos::TrimOligos(int p, int b, map<string, int> pr, map<string, int> br, v
                barcodes = br;
                primers = pr;
                revPrimer = r;
+        
+        maxFBarcodeLength = 0;
+        for(map<string,int>::iterator it=barcodes.begin();it!=barcodes.end();it++){
+            string oligo = it->first;
+            if(oligo.length() > maxFBarcodeLength){
+                maxFBarcodeLength = oligo.length();
+            }
+        }
+        maxRBarcodeLength = maxFBarcodeLength;
+        
+        maxFPrimerLength = 0;
+        for(map<string,int>::iterator it=primers.begin();it!=primers.end();it++){
+            string oligo = it->first;
+            if(oligo.length() > maxFPrimerLength){
+                maxFPrimerLength = oligo.length();
+            }
+        }
+        maxRPrimerLength = maxFPrimerLength;
        }
        catch(exception& e) {
                m->errorOut(e, "TrimOligos", "TrimOligos");
@@ -77,7 +170,135 @@ TrimOligos::TrimOligos(int p, int b, map<string, int> pr, map<string, int> br, v
 }
 /********************************************************************/
 TrimOligos::~TrimOligos() {}
+//********************************************************************/
+bool TrimOligos::findForward(Sequence& seq, int& primerStart, int& primerEnd){
+       try {
+               
+               string rawSequence = seq.getUnaligned();
+               
+               for(map<string,int>::iterator it=primers.begin();it!=primers.end();it++){
+                       string oligo = it->first;
+                       
+                       if(rawSequence.length() < oligo.length()) {  break;  }
+                       
+                       //search for primer
+            int olength = oligo.length();
+            for (int j = 0; j < rawSequence.length()-olength; j++){
+                if (m->control_pressed) {  primerStart = 0; primerEnd = 0; return false; }
+                string rawChunk = rawSequence.substr(j, olength);
+                if(compareDNASeq(oligo, rawChunk)) {
+                    primerStart = j;
+                    primerEnd = primerStart + olength;
+                    return true;
+                }
+                
+            }
+        }      
+               
+        primerStart = 0; primerEnd = 0;
+        //if you don't want to allow for diffs
+               if (pdiffs == 0) { return false;  }
+               else { //try aligning and see if you can find it
+                                               
+                       //can you find the barcode
+                       int minDiff = 1e6;
+                       int minCount = 1;
+                       
+            Alignment* alignment;
+            if (primers.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFPrimerLength+pdiffs+1));  }
+            else{ alignment = NULL; } 
+            
+                       for(map<string,int>::iterator it=primers.begin();it!=primers.end();it++){
+                
+                string prim = it->first;
+                //search for primer
+                int olength = prim.length();
+                if (rawSequence.length() < olength) {} //ignore primers too long for this seq
+                else{
+                    for (int j = 0; j < rawSequence.length()-olength; j++){
+                        
+                        string oligo = it->first;
+
+                        if (m->control_pressed) {  primerStart = 0; primerEnd = 0; return false; }
+                               
+                        string rawChunk = rawSequence.substr(j, olength+pdiffs);
+                        
+                        //use needleman to align first primer.length()+numdiffs of sequence to each barcode
+                        alignment->align(oligo, rawChunk);
+                        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 numDiff = countDiffs(oligo, temp);
+                               
+                        if(numDiff < minDiff){
+                            minDiff = numDiff;
+                            minCount = 1;
+                            primerStart = j;
+                            primerEnd = primerStart + alnLength;
+                        }else if(numDiff == minDiff){ minCount++; }
+                    }
+                }
+                       }
+                       
+            if (alignment != NULL) {  delete alignment;  }
+            
+                       if(minDiff > pdiffs)    {       primerStart = 0; primerEnd = 0; return false;           }       //no good matches
+                       else if(minCount > 1)   {       primerStart = 0; primerEnd = 0; return false;   }       //can't tell the difference between multiple primers
+                       else{  return true; }
+               }
+       
+        primerStart = 0; primerEnd = 0;
+               return false;
+               
+       }
+       catch(exception& e) {
+               m->errorOut(e, "TrimOligos", "stripForward");
+               exit(1);
+       }
+}
+//******************************************************************/
+bool TrimOligos::findReverse(Sequence& seq, int& primerStart, int& primerEnd){
+       try {
+               
+               string rawSequence = seq.getUnaligned();
+               
+               for(int i=0;i<revPrimer.size();i++){
+                       string oligo = revPrimer[i];
+                       if(rawSequence.length() < oligo.length()) {  break;  }
+                       
+                       //search for primer
+            int olength = oligo.length();
+            for (int j = rawSequence.length()-olength; j >= 0; j--){
+                if (m->control_pressed) {  primerStart = 0; primerEnd = 0; return false; }
+                string rawChunk = rawSequence.substr(j, olength);
+                
+                if(compareDNASeq(oligo, rawChunk)) {
+                    primerStart = j;
+                    primerEnd = primerStart + olength;
+                    return true;
+                }
+                
+            }
+               }       
+               
+        primerStart = 0; primerEnd = 0;
+               return false;
+       }
+       catch(exception& e) {
+               m->errorOut(e, "PcrSeqsCommand", "findReverse");
+               exit(1);
+       }
+}
 //*******************************************************************/
+
 int TrimOligos::stripBarcode(Sequence& seq, QualityScores& qual, int& group){
        try {
                
@@ -109,21 +330,9 @@ int TrimOligos::stripBarcode(Sequence& seq, QualityScores& qual, int& group){
                if ((bdiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (barcodes.size() > 0) {
-                               map<string,int>::iterator it; 
-                               
-                               for(it=barcodes.begin();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; } 
+                       if (barcodes.size() > 0) {alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFBarcodeLength+bdiffs+1));   }
+                       else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -135,7 +344,7 @@ int TrimOligos::stripBarcode(Sequence& seq, QualityScores& qual, int& group){
                                string oligo = it->first;
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                               if(rawSequence.length() < maxFBarcodeLength){   //let's just assume that the barcodes are the same length
                                        success = bdiffs + 10;
                                        break;
                                }
@@ -194,27 +403,33 @@ int TrimOligos::stripBarcode(Sequence& seq, QualityScores& qual, int& group){
                m->errorOut(e, "TrimOligos", "stripBarcode");
                exit(1);
        }
-       
 }
 //*******************************************************************/
-int TrimOligos::stripBarcode(Sequence& seq, int& group){
+int TrimOligos::stripBarcode(Sequence& forwardSeq, Sequence& reverseSeq, int& group){
        try {
-               
-               string rawSequence = seq.getUnaligned();
+               //look for forward barcode
+               string rawFSequence = forwardSeq.getUnaligned();
+        string rawRSequence = reverseSeq.getUnaligned();
                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
+               //can you find the forward barcode
+               for(map<int,oligosPair>::iterator it=ipbarcodes.begin();it!=ipbarcodes.end();it++){
+                       string foligo = it->second.forward;
+            string roligo = it->second.reverse;
+            
+                       if(rawFSequence.length() < foligo.length()){    //let's just assume that the barcodes are the same length
+                               success = bdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+            if(rawRSequence.length() < roligo.length()){       //let's just assume that the barcodes are the same length
                                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()));
-                               
+                       if((compareDNASeq(foligo, rawFSequence.substr(0,foligo.length()))) && (compareDNASeq(roligo, rawRSequence.substr((rawRSequence.length()-roligo.length()),roligo.length())))) {
+                               group = it->first;
+                               forwardSeq.setUnaligned(rawFSequence.substr(foligo.length()));
+                reverseSeq.setUnaligned(rawRSequence.substr(0,(rawRSequence.length()-roligo.length())));
                                success = 0;
                                break;
                        }
@@ -222,230 +437,899 @@ int TrimOligos::stripBarcode(Sequence& seq, int& group){
                
                //if you found the barcode or if you don't want to allow for diffs
                if ((bdiffs == 0) || (success == 0)) { return success;  }
-               
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (barcodes.size() > 0) {
-                               map<string,int>::iterator it=barcodes.begin();
-                               
-                               for(map<string,int>::iterator it=barcodes.begin();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; } 
+                       if (ifbarcodes.size() > 0) {  alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFBarcodeLength+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++){
+                       vector< vector<int> > minFGroup;
+                       vector<int> minFPos; 
+            
+            //the pair can have duplicates, but we only want to search for the unique forward and reverses, example
+            /*
+                1   Sarah   Westcott
+                2   John    Westcott
+                3   Anna    Westcott
+                4   Sarah   Schloss
+                5   Pat     Schloss
+                6   Gail    Brown
+                7   Pat     Moore
+             
+                only want to look for forward = Sarah, John, Anna, Pat, Gail
+                                      reverse = Westcott, Schloss, Brown, Moore
+             
+                but if best match forward = 4, and reverse = 1, we want to count as a valid match because forward 1 and forward 4 are the same.  so both barcodes map to same group.
+             */
+            //cout << endl << forwardSeq.getName() << endl;         
+                       for(map<string, vector<int> >::iterator it=ifbarcodes.begin();it!=ifbarcodes.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
+                               if(rawFSequence.length() < maxFBarcodeLength){  //let's just assume that the barcodes are the same length
                                        success = bdiffs + 10;
                                        break;
                                }
-                               
+                               //cout << "before = " << oligo << '\t' << rawFSequence.substr(0,oligo.length()+bdiffs) << endl;
                                //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
-                               alignment->align(oligo, rawSequence.substr(0,oligo.length()+bdiffs));
+                               alignment->align(oligo, rawFSequence.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;  }
-                               }
+                               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 numDiff = countDiffs(oligo, temp);
+                int numDiff = countDiffs(oligo, temp);
                                
+                if (alnLength == 0) { numDiff = bdiffs + 100; }
+                //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                
                                if(numDiff < minDiff){
                                        minDiff = numDiff;
                                        minCount = 1;
-                                       minGroup = it->second;
-                                       minPos = 0;
+                                       minFGroup.clear();
+                    minFGroup.push_back(it->second);
+                                       int tempminFPos = 0;
+                    minFPos.clear();
                                        for(int i=0;i<alnLength;i++){
                                                if(temp[i] != '-'){
-                                                       minPos++;
+                                                       tempminFPos++;
                                                }
                                        }
+                    minFPos.push_back(tempminFPos);
+                               }else if(numDiff == minDiff){
+                                       minFGroup.push_back(it->second);
+                    int tempminFPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
                                }
-                               else if(numDiff == minDiff){
-                                       minCount++;
-                               }
-                               
                        }
-                       
+            
+                       //cout << minDiff << '\t' << minCount << '\t' << endl;
                        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;
+                       else{   
+                //check for reverse match
+                if (alignment != NULL) {  delete alignment;  }
+                
+                if (irbarcodes.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxRBarcodeLength+bdiffs+1));   }
+                else{ alignment = NULL; } 
+                
+                //can you find the barcode
+                minDiff = 1e6;
+                minCount = 1;
+                vector< vector<int> > minRGroup;
+                vector<int> minRPos;
+                
+                for(map<string, vector<int> >::iterator it=irbarcodes.begin();it!=irbarcodes.end();it++){
+                    string oligo = it->first;
+                    //cout << "before = " << oligo << '\t' << rawRSequence.substr(0,oligo.length()+bdiffs) << endl;
+                    if(rawRSequence.length() < maxRBarcodeLength){     //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, rawRSequence.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 numDiff = countDiffs(oligo, temp);
+                    if (alnLength == 0) { numDiff = bdiffs + 100; }
+                    
+                    //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                    if(numDiff < minDiff){
+                        minDiff = numDiff;
+                        minCount = 1;
+                        minRGroup.clear();
+                        minRGroup.push_back(it->second);
+                        int tempminRPos = 0;
+                        minRPos.clear();
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);                    
+                    }else if(numDiff == minDiff){
+                        int tempminRPos = 0;
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);  
+                        minRGroup.push_back(it->second);
+                    }
+                    
+                }
+                
+                /*cout << minDiff << '\t' << minCount << '\t' << endl;
+                for (int i = 0; i < minFGroup.size(); i++) { 
+                    cout << i << '\t';
+                    for (int j = 0; j < minFGroup[i].size(); j++) {  cout << minFGroup[i][j] << " "; }
+                    cout << endl;
+                }
+                cout << endl;
+                for (int i = 0; i < minRGroup.size(); i++) { 
+                    cout << i << '\t';
+                    for (int j = 0; j < minRGroup[i].size(); j++) {  cout << minRGroup[i][j] << " "; }
+                    cout << endl;
+                }
+                cout << endl;*/
+                if(minDiff > bdiffs)   {       success = minDiff;              }       //no good matches
+                else  {  
+                    bool foundMatch = false;
+                    vector<int> matches;
+                    for (int i = 0; i < minFGroup.size(); i++) {
+                        for (int j = 0; j < minFGroup[i].size(); j++) {
+                            for (int k = 0; k < minRGroup.size(); k++) {
+                                if (m->inUsersGroups(minFGroup[i][j], minRGroup[k])) { matches.push_back(minFGroup[i][j]); k+= minRGroup.size(); }
+                            }
+                        }
+                    }
+                    
+                    int fStart = 0;
+                    int rStart = 0;
+                    if (matches.size() == 1) { 
+                        foundMatch = true;
+                        group = matches[0]; 
+                        fStart = minFPos[0];
+                        rStart = minRPos[0];
+                    }
+                    
+                    //we have an acceptable match for the forward and reverse, but do they match?
+                    if (foundMatch) {
+                        forwardSeq.setUnaligned(rawFSequence.substr(fStart));
+                        reverseSeq.setUnaligned(rawRSequence.substr(rStart));
+                        success = minDiff;
+                    }else { success = bdiffs + 100;    }
+                }
                        }
                        
                        if (alignment != NULL) {  delete alignment;  }
-                       
                }
                
                return success;
                
        }
        catch(exception& e) {
-               m->errorOut(e, "TrimOligos", "stripBarcode");
+               m->errorOut(e, "TrimOligos", "stripIBarcode");
                exit(1);
        }
        
 }
 //*******************************************************************/
-int TrimOligos::stripRBarcode(Sequence& seq, QualityScores& qual, int& group){
+int TrimOligos::stripBarcode(Sequence& forwardSeq, Sequence& reverseSeq, QualityScores& forwardQual, QualityScores& reverseQual, int& group){
        try {
-               
-               string rawSequence = seq.getUnaligned();
+               //look for forward barcode
+               string rawFSequence = forwardSeq.getUnaligned();
+        string rawRSequence = reverseSeq.getUnaligned();
                int success = bdiffs + 1;       //guilty until proven innocent
                
-               //can you find the barcode
-               for(map<string,int>::iterator it=rbarcodes.begin();it!=rbarcodes.end();it++){
-                       string oligo = it->first;
-                       if(rawSequence.length() < oligo.length()){      //let's just assume that the barcodes are the same length
+               //can you find the forward barcode
+               for(map<int,oligosPair>::iterator it=ipbarcodes.begin();it!=ipbarcodes.end();it++){
+                       string foligo = it->second.forward;
+            string roligo = it->second.reverse;
+            
+                       if(rawFSequence.length() < foligo.length()){    //let's just assume that the barcodes are the same length
+                               success = bdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+            if(rawRSequence.length() < roligo.length()){       //let's just assume that the barcodes are the same length
                                success = bdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
                                break;  
                        }
+                       
+                       if((compareDNASeq(foligo, rawFSequence.substr(0,foligo.length()))) && (compareDNASeq(roligo, rawRSequence.substr((rawRSequence.length()-roligo.length()),roligo.length())))) {
+                               group = it->first;
+                               forwardSeq.setUnaligned(rawFSequence.substr(foligo.length()));
+                reverseSeq.setUnaligned(rawRSequence.substr(0,(rawRSequence.length()-roligo.length())));
+                forwardQual.trimQScores(foligo.length(), -1);
+                reverseQual.trimQScores(-1, rawRSequence.length()-roligo.length());
+                               success = 0;
+                               break;
+                       }
+               }
+               
+               //if you found the barcode or if you don't want to allow for diffs
+               if ((bdiffs == 0) || (success == 0)) { return success;  }
+               else { //try aligning and see if you can find it
+                       Alignment* alignment;
+                       if (ifbarcodes.size() > 0) {  alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFBarcodeLength+bdiffs+1));   }
+                       else{ alignment = NULL; } 
+                       
+                       //can you find the barcode
+                       int minDiff = 1e6;
+                       int minCount = 1;
+                       vector< vector<int> > minFGroup;
+                       vector<int> minFPos; 
             
-                       if(compareDNASeq(oligo, rawSequence.substr((rawSequence.length()-oligo.length()),oligo.length()))){
-                               group = it->second;
-                               seq.setUnaligned(rawSequence.substr(0,(rawSequence.length()-oligo.length())));
+            //the pair can have duplicates, but we only want to search for the unique forward and reverses, example
+            /*
+             1   Sarah   Westcott
+             2   John    Westcott
+             3   Anna    Westcott
+             4   Sarah   Schloss
+             5   Pat     Schloss
+             6   Gail    Brown
+             7   Pat     Moore
+             
+             only want to look for forward = Sarah, John, Anna, Pat, Gail
+             reverse = Westcott, Schloss, Brown, Moore
+             
+             but if best match forward = 4, and reverse = 1, we want to count as a valid match because forward 1 and forward 4 are the same.  so both barcodes map to same group.
+             */
+            //cout << endl << forwardSeq.getName() << endl;         
+                       for(map<string, vector<int> >::iterator it=ifbarcodes.begin();it!=ifbarcodes.end();it++){
+                               string oligo = it->first;
                                
-                               if(qual.getName() != ""){
-                                       qual.trimQScores(-1, rawSequence.length()-oligo.length());
+                               if(rawFSequence.length() < maxFBarcodeLength){  //let's just assume that the barcodes are the same length
+                                       success = bdiffs + 10;
+                                       break;
                                }
+                               //cout << "before = " << oligo << '\t' << rawFSequence.substr(0,oligo.length()+bdiffs) << endl;
+                               //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                               alignment->align(oligo, rawFSequence.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 numDiff = countDiffs(oligo, temp);
                                
+                if (alnLength == 0) { numDiff = bdiffs + 100; }
+                //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                
+                               if(numDiff < minDiff){
+                                       minDiff = numDiff;
+                                       minCount = 1;
+                                       minFGroup.clear();
+                    minFGroup.push_back(it->second);
+                                       int tempminFPos = 0;
+                    minFPos.clear();
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
+                               }else if(numDiff == minDiff){
+                                       minFGroup.push_back(it->second);
+                    int tempminFPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
+                               }
+                       }
+            
+                       //cout << minDiff << '\t' << minCount << '\t' << endl;
+                       if(minDiff > bdiffs)    {       success = minDiff;              }       //no good matches
+                       else{   
+                //check for reverse match
+                if (alignment != NULL) {  delete alignment;  }
+                
+                if (irbarcodes.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxRBarcodeLength+bdiffs+1));   }
+                else{ alignment = NULL; } 
+                
+                //can you find the barcode
+                minDiff = 1e6;
+                minCount = 1;
+                vector< vector<int> > minRGroup;
+                vector<int> minRPos;
+                
+                for(map<string, vector<int> >::iterator it=irbarcodes.begin();it!=irbarcodes.end();it++){
+                    string oligo = it->first;
+                    //cout << "before = " << oligo << '\t' << rawRSequence.substr(0,oligo.length()+bdiffs) << endl;
+                    if(rawRSequence.length() < maxRBarcodeLength){     //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, rawRSequence.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 numDiff = countDiffs(oligo, temp);
+                    if (alnLength == 0) { numDiff = bdiffs + 100; }
+                    
+                    //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                    if(numDiff < minDiff){
+                        minDiff = numDiff;
+                        minCount = 1;
+                        minRGroup.clear();
+                        minRGroup.push_back(it->second);
+                        int tempminRPos = 0;
+                        minRPos.clear();
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);                    
+                    }else if(numDiff == minDiff){
+                        int tempminRPos = 0;
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);  
+                        minRGroup.push_back(it->second);
+                    }
+                    
+                }
+                
+                /*cout << minDiff << '\t' << minCount << '\t' << endl;
+                 for (int i = 0; i < minFGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minFGroup[i].size(); j++) {  cout << minFGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;
+                 for (int i = 0; i < minRGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minRGroup[i].size(); j++) {  cout << minRGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;*/
+                if(minDiff > bdiffs)   {       success = minDiff;              }       //no good matches
+                else  {  
+                    bool foundMatch = false;
+                    vector<int> matches;
+                    for (int i = 0; i < minFGroup.size(); i++) {
+                        for (int j = 0; j < minFGroup[i].size(); j++) {
+                            for (int k = 0; k < minRGroup.size(); k++) {
+                                if (m->inUsersGroups(minFGroup[i][j], minRGroup[k])) { matches.push_back(minFGroup[i][j]); k+= minRGroup.size(); }
+                            }
+                        }
+                    }
+                    
+                    int fStart = 0;
+                    int rStart = 0;
+                    if (matches.size() == 1) { 
+                        foundMatch = true;
+                        group = matches[0]; 
+                        fStart = minFPos[0];
+                        rStart = minRPos[0];
+                    }
+                    
+                    //we have an acceptable match for the forward and reverse, but do they match?
+                    if (foundMatch) {
+                        forwardSeq.setUnaligned(rawFSequence.substr(fStart));
+                        reverseSeq.setUnaligned(rawRSequence.substr(rStart));
+                        forwardQual.trimQScores(fStart, -1);
+                        reverseQual.trimQScores(rStart, -1);
+                        success = minDiff;
+                    }else { success = bdiffs + 100;    }
+                }
+                       }
+                       
+                       if (alignment != NULL) {  delete alignment;  }
+               }
+               
+               return success;
+               
+       }
+       catch(exception& e) {
+               m->errorOut(e, "TrimOligos", "stripIBarcode");
+               exit(1);
+       }
+       
+}
+//*******************************************************************/
+int TrimOligos::stripForward(Sequence& forwardSeq, Sequence& reverseSeq, QualityScores& forwardQual, QualityScores& reverseQual, int& group){
+       try {
+               //look for forward barcode
+               string rawFSequence = forwardSeq.getUnaligned();
+        string rawRSequence = reverseSeq.getUnaligned();
+               int success = pdiffs + 1;       //guilty until proven innocent
+               
+               //can you find the forward barcode
+               for(map<int,oligosPair>::iterator it=ipprimers.begin();it!=ipprimers.end();it++){
+                       string foligo = it->second.forward;
+            string roligo = it->second.reverse;
+            
+                       if(rawFSequence.length() < foligo.length()){    //let's just assume that the barcodes are the same length
+                               success = pdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+            if(rawRSequence.length() < roligo.length()){       //let's just assume that the barcodes are the same length
+                               success = pdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+                       
+                       if((compareDNASeq(foligo, rawFSequence.substr(0,foligo.length()))) && (compareDNASeq(roligo, rawRSequence.substr((rawRSequence.length()-roligo.length()),roligo.length())))) {
+                               group = it->first;
+                               forwardSeq.setUnaligned(rawFSequence.substr(foligo.length()));
+                reverseSeq.setUnaligned(rawRSequence.substr(0,(rawRSequence.length()-roligo.length())));
+                forwardQual.trimQScores(foligo.length(), -1);
+                reverseQual.trimQScores(-1, rawRSequence.length()-roligo.length());
                                success = 0;
                                break;
                        }
                }
                
                //if you found the barcode or if you don't want to allow for diffs
-               if ((bdiffs == 0) || (success == 0)) { return success;  }
-               
+               if ((pdiffs == 0) || (success == 0)) { return success;  }
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (rbarcodes.size() > 0) {
-                               map<string,int>::iterator it; 
+                       if (ifprimers.size() > 0) {  alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFPrimerLength+pdiffs+1));   }
+                       else{ alignment = NULL; } 
+                       
+                       //can you find the barcode
+                       int minDiff = 1e6;
+                       int minCount = 1;
+                       vector< vector<int> > minFGroup;
+                       vector<int> minFPos; 
+            
+            //the pair can have duplicates, but we only want to search for the unique forward and reverses, example
+            /*
+             1   Sarah   Westcott
+             2   John    Westcott
+             3   Anna    Westcott
+             4   Sarah   Schloss
+             5   Pat     Schloss
+             6   Gail    Brown
+             7   Pat     Moore
+             
+             only want to look for forward = Sarah, John, Anna, Pat, Gail
+             reverse = Westcott, Schloss, Brown, Moore
+             
+             but if best match forward = 4, and reverse = 1, we want to count as a valid match because forward 1 and forward 4 are the same.  so both barcodes map to same group.
+             */
+            //cout << endl << forwardSeq.getName() << endl;         
+                       for(map<string, vector<int> >::iterator it=ifprimers.begin();it!=ifprimers.end();it++){
+                               string oligo = it->first;
                                
-                               for(it=rbarcodes.begin();it!=rbarcodes.end();it++){
-                                       if(it->first.length() > maxLength){
-                                               maxLength = it->first.length();
-                                       }
+                               if(rawFSequence.length() < maxFPrimerLength){   //let's just assume that the barcodes are the same length
+                                       success = pdiffs + 10;
+                                       break;
                                }
-                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+bdiffs+1));  
+                               //cout << "before = " << oligo << '\t' << rawFSequence.substr(0,oligo.length()+pdiffs) << endl;
+                               //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                               alignment->align(oligo, rawFSequence.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 numDiff = countDiffs(oligo, temp);
                                
-                       }else{ alignment = NULL; } 
+                if (alnLength == 0) { numDiff = pdiffs + 100; }
+                //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                
+                               if(numDiff < minDiff){
+                                       minDiff = numDiff;
+                                       minCount = 1;
+                                       minFGroup.clear();
+                    minFGroup.push_back(it->second);
+                                       int tempminFPos = 0;
+                    minFPos.clear();
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
+                               }else if(numDiff == minDiff){
+                                       minFGroup.push_back(it->second);
+                    int tempminFPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
+                               }
+                       }
+            
+                       //cout << minDiff << '\t' << minCount << '\t' << endl;
+                       if(minDiff > pdiffs)    {       success = minDiff;              }       //no good matches
+                       else{   
+                //check for reverse match
+                if (alignment != NULL) {  delete alignment;  }
+                
+                if (irbarcodes.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxRPrimerLength+pdiffs+1));   }
+                else{ alignment = NULL; } 
+                
+                //can you find the barcode
+                minDiff = 1e6;
+                minCount = 1;
+                vector< vector<int> > minRGroup;
+                vector<int> minRPos;
+                
+                for(map<string, vector<int> >::iterator it=irprimers.begin();it!=irprimers.end();it++){
+                    string oligo = it->first;
+                    //cout << "before = " << oligo << '\t' << rawRSequence.substr(0,oligo.length()+pdiffs) << endl;
+                    if(rawRSequence.length() < maxRPrimerLength){      //let's just assume that the barcodes are the same length
+                        success = pdiffs + 10;
+                        break;
+                    }
+                    
+                    //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                    alignment->align(oligo, rawRSequence.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 numDiff = countDiffs(oligo, temp);
+                    if (alnLength == 0) { numDiff = pdiffs + 100; }
+                    
+                    //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                    if(numDiff < minDiff){
+                        minDiff = numDiff;
+                        minCount = 1;
+                        minRGroup.clear();
+                        minRGroup.push_back(it->second);
+                        int tempminRPos = 0;
+                        minRPos.clear();
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);                    
+                    }else if(numDiff == minDiff){
+                        int tempminRPos = 0;
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);  
+                        minRGroup.push_back(it->second);
+                    }
+                    
+                }
+                
+                /*cout << minDiff << '\t' << minCount << '\t' << endl;
+                 for (int i = 0; i < minFGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minFGroup[i].size(); j++) {  cout << minFGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;
+                 for (int i = 0; i < minRGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minRGroup[i].size(); j++) {  cout << minRGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;*/
+                if(minDiff > pdiffs)   {       success = minDiff;              }       //no good matches
+                else  {  
+                    bool foundMatch = false;
+                    vector<int> matches;
+                    for (int i = 0; i < minFGroup.size(); i++) {
+                        for (int j = 0; j < minFGroup[i].size(); j++) {
+                            for (int k = 0; k < minRGroup.size(); k++) {
+                                if (m->inUsersGroups(minFGroup[i][j], minRGroup[k])) { matches.push_back(minFGroup[i][j]); k+= minRGroup.size(); }
+                            }
+                        }
+                    }
+                    
+                    int fStart = 0;
+                    int rStart = 0;
+                    if (matches.size() == 1) { 
+                        foundMatch = true;
+                        group = matches[0]; 
+                        fStart = minFPos[0];
+                        rStart = minRPos[0];
+                    }
+                    
+                    //we have an acceptable match for the forward and reverse, but do they match?
+                    if (foundMatch) {
+                        forwardSeq.setUnaligned(rawFSequence.substr(fStart));
+                        reverseSeq.setUnaligned(rawRSequence.substr(rStart));
+                        forwardQual.trimQScores(fStart, -1);
+                        reverseQual.trimQScores(rStart, -1);
+                        success = minDiff;
+                    }else { success = pdiffs + 100;    }
+                }
+                       }
+                       
+                       if (alignment != NULL) {  delete alignment;  }
+               }
+               
+               return success;
+               
+       }
+       catch(exception& e) {
+               m->errorOut(e, "TrimOligos", "stripIForward");
+               exit(1);
+       }
+       
+}
+//*******************************************************************/
+int TrimOligos::stripForward(Sequence& forwardSeq, Sequence& reverseSeq, int& group){
+       try {
+               //look for forward barcode
+               string rawFSequence = forwardSeq.getUnaligned();
+        string rawRSequence = reverseSeq.getUnaligned();
+               int success = pdiffs + 1;       //guilty until proven innocent
+               
+               //can you find the forward barcode
+               for(map<int,oligosPair>::iterator it=ipprimers.begin();it!=ipprimers.end();it++){
+                       string foligo = it->second.forward;
+            string roligo = it->second.reverse;
+            
+                       if(rawFSequence.length() < foligo.length()){    //let's just assume that the barcodes are the same length
+                               success = pdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+            if(rawRSequence.length() < roligo.length()){       //let's just assume that the barcodes are the same length
+                               success = pdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
+                               break;  
+                       }
+                       
+                       if((compareDNASeq(foligo, rawFSequence.substr(0,foligo.length()))) && (compareDNASeq(roligo, rawRSequence.substr((rawRSequence.length()-roligo.length()),roligo.length())))) {
+                               group = it->first;
+                               forwardSeq.setUnaligned(rawFSequence.substr(foligo.length()));
+                reverseSeq.setUnaligned(rawRSequence.substr(0,(rawRSequence.length()-roligo.length())));
+                               success = 0;
+                               break;
+                       }
+               }
+               
+               //if you found the barcode or if you don't want to allow for diffs
+               if ((pdiffs == 0) || (success == 0)) { return success;  }
+               else { //try aligning and see if you can find it
+                       Alignment* alignment;
+                       if (ifprimers.size() > 0) {  alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFPrimerLength+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=rbarcodes.begin();it!=rbarcodes.end();it++){
+                       vector< vector<int> > minFGroup;
+                       vector<int> minFPos; 
+            
+            //the pair can have duplicates, but we only want to search for the unique forward and reverses, example
+            /*
+             1   Sarah   Westcott
+             2   John    Westcott
+             3   Anna    Westcott
+             4   Sarah   Schloss
+             5   Pat     Schloss
+             6   Gail    Brown
+             7   Pat     Moore
+             
+             only want to look for forward = Sarah, John, Anna, Pat, Gail
+             reverse = Westcott, Schloss, Brown, Moore
+             
+             but if best match forward = 4, and reverse = 1, we want to count as a valid match because forward 1 and forward 4 are the same.  so both barcodes map to same group.
+             */
+            //cout << endl << forwardSeq.getName() << endl;         
+                       for(map<string, vector<int> >::iterator it=ifprimers.begin();it!=ifprimers.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;
+                               if(rawFSequence.length() < maxFPrimerLength){   //let's just assume that the barcodes are the same length
+                                       success = pdiffs + 10;
                                        break;
                                }
-                               
+                               //cout << "before = " << oligo << '\t' << rawFSequence.substr(0,oligo.length()+pdiffs) << endl;
                                //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
-                               alignment->align(oligo, rawSequence.substr((rawSequence.length()-(oligo.length()+bdiffs)),oligo.length()+bdiffs));
+                               alignment->align(oligo, rawFSequence.substr(0,oligo.length()+pdiffs));
                                oligo = alignment->getSeqAAln();
                                string temp = alignment->getSeqBAln();
-     
+                
                                int alnLength = oligo.length();
                                
-                               for(int i=0;i<alnLength;i++){
-                                       if(oligo[i] != '-'){    alnLength = i;  break;  }
-                               }
-                               oligo = oligo.substr(alnLength);
-                               temp = temp.substr(alnLength);
-                               
-                               int numDiff = countDiffs(oligo, temp);
+                               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 numDiff = countDiffs(oligo, temp);
                                
+                if (alnLength == 0) { numDiff = pdiffs + 100; }
+                //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                
                                if(numDiff < minDiff){
                                        minDiff = numDiff;
                                        minCount = 1;
-                                       minGroup = it->second;
-                                       minPos = 0;
-                                       for(int i=alnLength-1;i>=0;i--){
+                                       minFGroup.clear();
+                    minFGroup.push_back(it->second);
+                                       int tempminFPos = 0;
+                    minFPos.clear();
+                                       for(int i=0;i<alnLength;i++){
                                                if(temp[i] != '-'){
-                                                       minPos++;
+                                                       tempminFPos++;
                                                }
                                        }
+                    minFPos.push_back(tempminFPos);
+                               }else if(numDiff == minDiff){
+                                       minFGroup.push_back(it->second);
+                    int tempminFPos = 0;
+                                       for(int i=0;i<alnLength;i++){
+                                               if(temp[i] != '-'){
+                                                       tempminFPos++;
+                                               }
+                                       }
+                    minFPos.push_back(tempminFPos);
                                }
-                               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(0, (rawSequence.length()-minPos)));
+            
+                       //cout << minDiff << '\t' << minCount << '\t' << endl;
+                       if(minDiff > pdiffs)    {       success = minDiff;              }       //no good matches
+                       else{   
+                //check for reverse match
+                if (alignment != NULL) {  delete alignment;  }
                 
-                               if(qual.getName() != ""){
-                                       qual.trimQScores(-1, (rawSequence.length()-minPos));
-                               }
-                               success = minDiff;
+                if (irbarcodes.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxRPrimerLength+pdiffs+1));   }
+                else{ alignment = NULL; } 
+                
+                //can you find the barcode
+                minDiff = 1e6;
+                minCount = 1;
+                vector< vector<int> > minRGroup;
+                vector<int> minRPos;
+                
+                for(map<string, vector<int> >::iterator it=irprimers.begin();it!=irprimers.end();it++){
+                    string oligo = it->first;
+                    //cout << "before = " << oligo << '\t' << rawRSequence.substr(0,oligo.length()+pdiffs) << endl;
+                    if(rawRSequence.length() < maxRPrimerLength){      //let's just assume that the barcodes are the same length
+                        success = pdiffs + 10;
+                        break;
+                    }
+                    
+                    //use needleman to align first barcode.length()+numdiffs of sequence to each barcode
+                    alignment->align(oligo, rawRSequence.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 numDiff = countDiffs(oligo, temp);
+                    if (alnLength == 0) { numDiff = pdiffs + 100; }
+                    
+                    //cout << "after = " << oligo << '\t' << temp << '\t' << numDiff << endl;
+                    if(numDiff < minDiff){
+                        minDiff = numDiff;
+                        minCount = 1;
+                        minRGroup.clear();
+                        minRGroup.push_back(it->second);
+                        int tempminRPos = 0;
+                        minRPos.clear();
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);                    
+                    }else if(numDiff == minDiff){
+                        int tempminRPos = 0;
+                        for(int i=0;i<alnLength;i++){
+                            if(temp[i] != '-'){
+                                tempminRPos++;
+                            }
+                        }
+                        minRPos.push_back(tempminRPos);  
+                        minRGroup.push_back(it->second);
+                    }
+                    
+                }
+                
+                /*cout << minDiff << '\t' << minCount << '\t' << endl;
+                 for (int i = 0; i < minFGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minFGroup[i].size(); j++) {  cout << minFGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;
+                 for (int i = 0; i < minRGroup.size(); i++) { 
+                 cout << i << '\t';
+                 for (int j = 0; j < minRGroup[i].size(); j++) {  cout << minRGroup[i][j] << " "; }
+                 cout << endl;
+                 }
+                 cout << endl;*/
+                if(minDiff > pdiffs)   {       success = minDiff;              }       //no good matches
+                else  {  
+                    bool foundMatch = false;
+                    vector<int> matches;
+                    for (int i = 0; i < minFGroup.size(); i++) {
+                        for (int j = 0; j < minFGroup[i].size(); j++) {
+                            for (int k = 0; k < minRGroup.size(); k++) {
+                                if (m->inUsersGroups(minFGroup[i][j], minRGroup[k])) { matches.push_back(minFGroup[i][j]); k+= minRGroup.size(); }
+                            }
+                        }
+                    }
+                    
+                    int fStart = 0;
+                    int rStart = 0;
+                    if (matches.size() == 1) { 
+                        foundMatch = true;
+                        group = matches[0]; 
+                        fStart = minFPos[0];
+                        rStart = minRPos[0];
+                    }
+                    
+                    //we have an acceptable match for the forward and reverse, but do they match?
+                    if (foundMatch) {
+                        forwardSeq.setUnaligned(rawFSequence.substr(fStart));
+                        reverseSeq.setUnaligned(rawRSequence.substr(rStart));
+                        success = minDiff;
+                    }else { success = pdiffs + 100;    }
+                }
                        }
                        
                        if (alignment != NULL) {  delete alignment;  }
-                       
                }
                
                return success;
                
        }
        catch(exception& e) {
-               m->errorOut(e, "TrimOligos", "stripRBarcode");
+               m->errorOut(e, "TrimOligos", "stripIForward");
                exit(1);
        }
        
 }
 //*******************************************************************/
-int TrimOligos::stripRBarcode(Sequence& seq, int& group){
+int TrimOligos::stripBarcode(Sequence& seq, int& group){
        try {
                
                string rawSequence = seq.getUnaligned();
                int success = bdiffs + 1;       //guilty until proven innocent
                
                //can you find the barcode
-               for(map<string,int>::iterator it=rbarcodes.begin();it!=rbarcodes.end();it++){
+               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 = bdiffs + 10;                                  //if the sequence is shorter than the barcode then bail out
                                break;  
                        }
-            
-                       if(compareDNASeq(oligo, rawSequence.substr((rawSequence.length()-oligo.length()),oligo.length()))){
+                       
+                       if(compareDNASeq(oligo, rawSequence.substr(0,oligo.length()))){
                                group = it->second;
-                               seq.setUnaligned(rawSequence.substr(0,(rawSequence.length()-oligo.length())));
+                               seq.setUnaligned(rawSequence.substr(oligo.length()));
                                
                                success = 0;
                                break;
@@ -456,21 +1340,9 @@ int TrimOligos::stripRBarcode(Sequence& seq, int& group){
                if ((bdiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
-                       Alignment* alignment;
-                       if (rbarcodes.size() > 0) {
-                               map<string,int>::iterator it; 
-                               
-                               for(it=rbarcodes.begin();it!=rbarcodes.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; } 
+            Alignment* alignment;
+                       if (barcodes.size() > 0) {  alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFBarcodeLength+bdiffs+1));  }
+                       else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -478,28 +1350,28 @@ int TrimOligos::stripRBarcode(Sequence& seq, int& group){
                        int minGroup = -1;
                        int minPos = 0;
                        
-                       for(map<string,int>::iterator it=rbarcodes.begin();it!=rbarcodes.end();it++){
+                       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
+                               if(rawSequence.length() < maxFBarcodeLength){   //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((rawSequence.length()-(oligo.length()+bdiffs)),oligo.length()+bdiffs));
+                               alignment->align(oligo, rawSequence.substr(0,oligo.length()+bdiffs));
                                oligo = alignment->getSeqAAln();
                                string temp = alignment->getSeqBAln();
-                
+                                
                                int alnLength = oligo.length();
                                
-                               for(int i=0;i<alnLength;i++){
-                                       if(oligo[i] != '-'){    alnLength = i;  break;  }
+                               for(int i=oligo.length()-1;i>=0;i--){
+                                       if(oligo[i] != '-'){    alnLength = i+1;        break;  }
                                }
-                               oligo = oligo.substr(alnLength);
-                               temp = temp.substr(alnLength);
-                               
+                               oligo = oligo.substr(0,alnLength);
+                               temp = temp.substr(0,alnLength);
+                                
                                int numDiff = countDiffs(oligo, temp);
                                
                                if(numDiff < minDiff){
@@ -507,7 +1379,7 @@ int TrimOligos::stripRBarcode(Sequence& seq, int& group){
                                        minCount = 1;
                                        minGroup = it->second;
                                        minPos = 0;
-                                       for(int i=alnLength-1;i>=0;i--){
+                                       for(int i=0;i<alnLength;i++){
                                                if(temp[i] != '-'){
                                                        minPos++;
                                                }
@@ -523,8 +1395,7 @@ int TrimOligos::stripRBarcode(Sequence& seq, int& group){
                        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(0, (rawSequence.length()-minPos)));
-                
+                               seq.setUnaligned(rawSequence.substr(minPos));
                                success = minDiff;
                        }
                        
@@ -536,12 +1407,13 @@ int TrimOligos::stripRBarcode(Sequence& seq, int& group){
                
        }
        catch(exception& e) {
-               m->errorOut(e, "TrimOligos", "stripRBarcode");
+               m->errorOut(e, "TrimOligos", "stripBarcode");
                exit(1);
        }
        
 }
-//********************************************************************/
+
+/********************************************************************/
 int TrimOligos::stripForward(Sequence& seq, int& group){
        try {
                
@@ -568,21 +1440,9 @@ int TrimOligos::stripForward(Sequence& seq, int& group){
                if ((pdiffs == 0) || (success == 0)) {  return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (primers.size() > 0) {
-                               map<string,int>::iterator it; 
-                               
-                               for(it=primers.begin();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; } 
+                       if (primers.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFPrimerLength+pdiffs+1));   }
+            else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -594,7 +1454,7 @@ int TrimOligos::stripForward(Sequence& seq, int& group){
                                string oligo = it->first;
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   
+                               if(rawSequence.length() < maxFPrimerLength){    
                                        success = pdiffs + 100;
                                        break;
                                }
@@ -680,21 +1540,9 @@ int TrimOligos::stripForward(Sequence& seq, QualityScores& qual, int& group, boo
                if ((pdiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (primers.size() > 0) {
-                               map<string,int>::iterator it; 
-                               
-                               for(it=primers.begin();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; } 
+                       if (primers.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxFPrimerLength+pdiffs+1));  }
+                       else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -706,7 +1554,7 @@ int TrimOligos::stripForward(Sequence& seq, QualityScores& qual, int& group, boo
                                string oligo = it->first;
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   
+                               if(rawSequence.length() < maxFPrimerLength){    
                                        success = pdiffs + 100;
                                        break;
                                }
@@ -855,19 +1703,9 @@ bool TrimOligos::stripLinker(Sequence& seq, QualityScores& qual){
                if ((ldiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (linker.size() > 0) {
-                               for(int i = 0; i < linker.size(); i++){
-                                       if(linker[i].length() > maxLength){
-                                               maxLength = linker[i].length();
-                                       }
-                               }
-                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+ldiffs+1));  
-                               
-                       }else{ alignment = NULL; } 
+                       if (linker.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLinkerLength+ldiffs+1));   }     
+                       else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -878,7 +1716,7 @@ bool TrimOligos::stripLinker(Sequence& seq, QualityScores& qual){
                                string oligo = linker[i];
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                               if(rawSequence.length() < maxLinkerLength){     //let's just assume that the barcodes are the same length
                                        success = ldiffs + 10;
                                        break;
                                }
@@ -964,19 +1802,9 @@ bool TrimOligos::stripLinker(Sequence& seq){
                if ((ldiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (linker.size() > 0) {
-                               for(int i = 0; i < linker.size(); i++){
-                                       if(linker[i].length() > maxLength){
-                                               maxLength = linker[i].length();
-                                       }
-                               }
-                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+ldiffs+1));  
-                               
-                       }else{ alignment = NULL; } 
+                       if (linker.size() > 0) {alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLinkerLength+ldiffs+1));  }
+            else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -987,7 +1815,7 @@ bool TrimOligos::stripLinker(Sequence& seq){
                                string oligo = linker[i];
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                               if(rawSequence.length() < maxLinkerLength){     //let's just assume that the barcodes are the same length
                                        success = ldiffs + 10;
                                        break;
                                }
@@ -1071,19 +1899,9 @@ bool TrimOligos::stripSpacer(Sequence& seq, QualityScores& qual){
                if ((sdiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (spacer.size() > 0) {
-                               for(int i = 0; i < spacer.size(); i++){
-                                       if(spacer[i].length() > maxLength){
-                                               maxLength = spacer[i].length();
-                                       }
-                               }
-                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+sdiffs+1));  
-                               
-                       }else{ alignment = NULL; } 
+                       if (spacer.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxSpacerLength+sdiffs+1));  }
+            else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -1094,7 +1912,7 @@ bool TrimOligos::stripSpacer(Sequence& seq, QualityScores& qual){
                                string oligo = spacer[i];
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                               if(rawSequence.length() < maxSpacerLength){     //let's just assume that the barcodes are the same length
                                        success = sdiffs + 10;
                                        break;
                                }
@@ -1180,19 +1998,9 @@ bool TrimOligos::stripSpacer(Sequence& seq){
                if ((sdiffs == 0) || (success == 0)) { return success;  }
                
                else { //try aligning and see if you can find it
-                       
-                       int maxLength = 0;
-                       
                        Alignment* alignment;
-                       if (spacer.size() > 0) {
-                               for(int i = 0; i < spacer.size(); i++){
-                                       if(spacer[i].length() > maxLength){
-                                               maxLength = spacer[i].length();
-                                       }
-                               }
-                               alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxLength+sdiffs+1));  
-                               
-                       }else{ alignment = NULL; } 
+                       if (spacer.size() > 0) { alignment = new NeedlemanOverlap(-1.0, 1.0, -1.0, (maxSpacerLength+sdiffs+1));  }
+            else{ alignment = NULL; } 
                        
                        //can you find the barcode
                        int minDiff = 1e6;
@@ -1203,7 +2011,7 @@ bool TrimOligos::stripSpacer(Sequence& seq){
                                string oligo = spacer[i];
                                //                              int length = oligo.length();
                                
-                               if(rawSequence.length() < maxLength){   //let's just assume that the barcodes are the same length
+                               if(rawSequence.length() < maxSpacerLength){     //let's just assume that the barcodes are the same length
                                        success = sdiffs + 10;
                                        break;
                                }