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#include "ChimericAlign.h"
void ChimericAlign::chimericStitching(char *genSeq, char **Read1) {
if (stitchingDone)
return;
stitchingDone=true;
char *readSeq=Read1[0]; //only direct read sequence is used - reverse complemented if necessary in the algorithm
al1=new Transcript(*al1);
al2=new Transcript(*al2);
Transcript &a1=*al1;
Transcript &a2=*al2;//to use instead of pointers
chimStr = max(seg1.str,seg2.str); //segment strands are either equal, or one is zero - select the non-zero strand
chimRepeat1=0; chimRepeat2=0; chimJ1=0; chimJ2=0; chimMotif=0;
if ( a1.exons[ex1][EX_iFrag] < a2.exons[ex2][EX_iFrag] ) {//mates bracket the chimeric junction
chimMotif=-1;
if (a1.Str==1) {//negative strand
chimJ1=a1.exons[ex1][EX_G]-1;
} else {
chimJ1=a1.exons[ex1][EX_G]+a1.exons[ex1][EX_L];
};
if (a2.Str==0) {//positive strand
chimJ2=a2.exons[ex2][EX_G]-1;
} else {
chimJ2=a2.exons[ex2][EX_G]+a2.exons[ex2][EX_L];
};
} else {//chimeric junctions is within one of the mates, check and shift chimeric junction if necessary
uint roStart0 = a1.Str==0 ? a1.exons[ex1][EX_R] : a1.Lread - a1.exons[ex1][EX_R] - a1.exons[ex1][EX_L];
uint roStart1 = a2.Str==0 ? a2.exons[ex2][EX_R] : a1.Lread - a2.exons[ex2][EX_R] - a2.exons[ex2][EX_L];
uint jR, jRbest=0;
int jScore=0,jMotif=0,jScoreBest=-999999,jScoreJ=0;
uint jRmax = roStart1+a2.exons[ex2][EX_L];
jRmax = jRmax>roStart0 ? jRmax-roStart0-1 : 0;
for (jR=0; jR<jRmax; jR++) {//scan through the exons to find a canonical junction, and check for mismatches
if (jR==a1.readLength[0]) jR++; //skip the inter-mate base
char bR=readSeq[roStart0+jR];
char b0,b1;
if (a1.Str==0) {
b0=genSeq[a1.exons[ex1][EX_G]+jR];
} else {
b0=genSeq[a1.exons[ex1][EX_G]+a1.exons[ex1][EX_L]-1-jR];
if (b0<4) b0=3-b0;
};
if (a2.Str==0) {
b1=genSeq[a2.exons[ex2][EX_G]-roStart1+roStart0+jR];
} else {
b1=genSeq[a2.exons[ex2][EX_G]+a2.exons[ex2][EX_L]-1+roStart1-roStart0-jR];
if (b1<4) b1=3-b1;
};
if ( ( P.pCh.filter.genomicN && (b0>3 || b1>3) ) || bR>3) {//chimera is not called if there are Ns in the genome or in the read
chimScore=0;
return;
};
char b01,b02,b11,b12;
if (a1.Str==0) {
b01=genSeq[a1.exons[ex1][EX_G]+jR+1];
b02=genSeq[a1.exons[ex1][EX_G]+jR+2];
} else {
b01=genSeq[a1.exons[ex1][EX_G]+a1.exons[ex1][EX_L]-1-jR-1];
if (b01<4) b01=3-b01;
b02=genSeq[a1.exons[ex1][EX_G]+a1.exons[ex1][EX_L]-1-jR-2];
if (b02<4) b02=3-b02;
};
if (a2.Str==0) {
b11=genSeq[a2.exons[ex2][EX_G]-roStart1+roStart0+jR-1];
b12=genSeq[a2.exons[ex2][EX_G]-roStart1+roStart0+jR];
} else {
b11=genSeq[a2.exons[ex2][EX_G]+a2.exons[ex2][EX_L]-1+roStart1-roStart0-jR+1];
if (b11<4) b11=3-b11;
b12=genSeq[a2.exons[ex2][EX_G]+a2.exons[ex2][EX_L]-1+roStart1-roStart0-jR];
if (b12<4) b12=3-b12;
};
jMotif=0;
if (b01==2 && b02==3 && b11==0 && b12==2) {//GTAG
if (chimStr!=2) {
jMotif=1;
};
} else if(b01==1 && b02==3 && b11==0 && b12==1) {//CTAC
if (chimStr!=1) {
jMotif=2;
};
};
if (bR==b0 && bR!=b1) {
jScore++;
} else if (bR!=b0 && bR==b1) {
jScore--;
};
jScoreJ =jMotif==0 ? jScore + P.pCh.scoreJunctionNonGTAG : jScore ;
if ( jScoreJ > jScoreBest || (jScoreJ == jScoreBest && jMotif>0) ) {
chimMotif=jMotif;
jRbest=jR;
jScoreBest=jScoreJ;
};
};//jR cycle
//shift junction in trChim
if (a1.Str==1) {
a1.exons[ex1][EX_R] +=a1.exons[ex1][EX_L]-jRbest-1;
a1.exons[ex1][EX_G] +=a1.exons[ex1][EX_L]-jRbest-1;
a1.exons[ex1][EX_L]=jRbest+1;
chimJ1=a1.exons[ex1][EX_G]-1;
} else {
a1.exons[ex1][EX_L]=jRbest+1;
chimJ1=a1.exons[ex1][EX_G]+a1.exons[ex1][EX_L];
};
if (a2.Str==0) {
a2.exons[ex2][EX_R] +=roStart0+jRbest+1-roStart1;
a2.exons[ex2][EX_G] +=roStart0+jRbest+1-roStart1;
a2.exons[ex2][EX_L]=roStart1+a2.exons[ex2][EX_L]-roStart0-jRbest-1;
chimJ2=a2.exons[ex2][EX_G]-1;
} else {
a2.exons[ex2][EX_L]=roStart1+a2.exons[ex2][EX_L]-roStart0-jRbest-1;
chimJ2=a2.exons[ex2][EX_G]+a2.exons[ex2][EX_L];
};
//find repeats
char b0,b1;
for (jR=0;jR<100;jR++) {//forward check
if (a1.Str==0) {
b0=genSeq[chimJ1+jR];
} else {
b0=genSeq[chimJ1-jR];
if (b0<4) b0=3-b0;
};
if (a2.Str==0) {
b1=genSeq[chimJ2+1+jR];
} else {
b1=genSeq[chimJ2-1-jR];
if (b1<4) b1=3-b1;
};
if (b0!=b1) break;
};
chimRepeat2=jR;
for (jR=0;jR<100;jR++) {//reverse check
if (a1.Str==0) {
b0=genSeq[chimJ1-1-jR];
} else {
b0=genSeq[chimJ1+1+jR];
if (b0<4) b0=3-b0;
};
if (a2.Str==0) {
b1=genSeq[chimJ2-jR];
} else {
b1=genSeq[chimJ2+jR];
if (b1<4) b1=3-b1;
};
if (b0!=b1) break;
};
chimRepeat1=jR;
};//chimeric junction is within a mate
if (chimMotif>=0 && (a1.exons[ex1][EX_L]<P.pCh.junctionOverhangMin || a2.exons[ex2][EX_L]<P.pCh.junctionOverhangMin) ) {
//filter out cases where linear junctions that are very close to chimeric junction
chimScore=0;
return;
};
//re-calculate chimScore for adjusted transcripts
chimScore=a1.alignScore(Read1,genSeq,P) + a2.alignScore(Read1,genSeq,P) + (chimMotif==0 ? P.pCh.scoreJunctionNonGTAG : 0);
};
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