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sam_funcs.c
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sam_funcs.c
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#include "sam_funcs.h"
#undef __MY_DEBUG__
#undef __MY_DEBUG_READ__
#ifdef __MY_DEBUG__
#define __DEBUG_CHR__ "chr11"
#define __DEBUG_POS__ 75279846
#endif
#define WARNING_THR 2147483647
void snpAnalysis(char* bamFileName, char* snpFileName, char* methCgFileName, char* methChgFileName, char* methChhFileName, HashNode** hashTable, char** chrSeqArray, int* chrLen, int chrCnt, int vQualMin, int nLayerMin, int nLayerMax, float vSnpRate, float vSnpPerBase, unsigned int mapqThr)
{
methProcess(bamFileName, methCgFileName, methChgFileName, methChhFileName, hashTable, chrSeqArray, chrLen, chrCnt, vQualMin, nLayerMin, mapqThr);
snpProcess(bamFileName, snpFileName, hashTable, chrSeqArray, chrLen, chrCnt, vQualMin, nLayerMax, vSnpRate, vSnpPerBase, mapqThr);
}
void printMeth(FILE* methFptr, int len, char* curChr, unsigned int* w_Mm_CG, unsigned int* w_Mc_CG, unsigned int* w_Mq_CG, unsigned int* c_Mm_CG, unsigned int* c_Mc_CG, unsigned int* c_Mq_CG, const char* tag, int nLayerMin)
{
int i;
// Record meth sites
if(strcmp(tag, "CG") == 0) {
for(i = 0; i < len - 1; i++) {
if(w_Mc_CG[i] + c_Mc_CG[i+1] >= nLayerMin)
if(w_Mc_CG[i] != 0 && c_Mc_CG[i+1] != 0)
fprintf(methFptr, "%s\t%d\t%s\t%d\t%d\t%d\t%d\t%d\t%d\n", curChr, i + 1, tag, w_Mm_CG[i], w_Mc_CG[i], (unsigned int)(w_Mq_CG[i]/w_Mc_CG[i]), c_Mm_CG[i+1], c_Mc_CG[i+1], (unsigned int)(c_Mq_CG[i+1]/c_Mc_CG[i+1]));
else if(w_Mc_CG[i] == 0)
fprintf(methFptr, "%s\t%d\t%s\t.\t.\t.\t%d\t%d\t%d\n", curChr, i + 1, tag, c_Mm_CG[i+1], c_Mc_CG[i+1], (unsigned int)(c_Mq_CG[i+1]/c_Mc_CG[i+1]));
else
fprintf(methFptr, "%s\t%d\t%s\t%d\t%d\t%d\t.\t.\t.\n", curChr, i + 1, tag, w_Mm_CG[i], w_Mc_CG[i], (unsigned int)(w_Mq_CG[i]/w_Mc_CG[i]));
}
}
else if(strcmp(tag, "CHG") == 0){
for(i = 0; i < len - 2; i++) {
if(w_Mc_CG[i] + c_Mc_CG[i+2] >= nLayerMin)
if(w_Mc_CG[i] != 0 && c_Mc_CG[i+2] != 0)
fprintf(methFptr, "%s\t%d\t%s\t%d\t%d\t%d\t%d\t%d\t%d\n", curChr, i + 1, tag, w_Mm_CG[i], w_Mc_CG[i], (unsigned int)(w_Mq_CG[i]/w_Mc_CG[i]), c_Mm_CG[i+2], c_Mc_CG[i+2], (unsigned int)(c_Mq_CG[i+2]/c_Mc_CG[i+2]));
else if(w_Mc_CG[i] == 0)
fprintf(methFptr, "%s\t%d\t%s\t.\t.\t.\t%d\t%d\t%d\n", curChr, i + 3, tag, c_Mm_CG[i+2], c_Mc_CG[i+2], (unsigned int)(c_Mq_CG[i+2]/c_Mc_CG[i+2]));
else
fprintf(methFptr, "%s\t%d\t%s\t%d\t%d\t%d\t.\t.\t.\n", curChr, i + 1, tag, w_Mm_CG[i], w_Mc_CG[i], (unsigned int)(w_Mq_CG[i]/w_Mc_CG[i]));
}
}
else {
for(i = 0; i < len - 2; i++) {
if(w_Mc_CG[i] + c_Mc_CG[i+2] >= nLayerMin) {
if(w_Mc_CG[i] > 0)
fprintf(methFptr, "%s\t%d\t%s\t%d\t%d\t%d\t.\t.\t.\n", curChr, i + 1, tag, w_Mm_CG[i], w_Mc_CG[i], (unsigned int)(w_Mq_CG[i]/w_Mc_CG[i]));
if(c_Mc_CG[i+2] > 0)
fprintf(methFptr, "%s\t%d\t%s\t.\t.\t.\t%d\t%d\t%d\n", curChr, i + 3, tag, c_Mm_CG[i+2], c_Mc_CG[i+2], (unsigned int)(c_Mq_CG[i+2]/c_Mc_CG[i+2]));
}
}
}
}
void printSnp(FILE* posFptr, char** chrSeqArray, int idx, int len, float vSnpRate, char* curChr, unsigned int *w_A, unsigned int *w_T, unsigned int *w_C, unsigned int *w_G, unsigned int *c_A, unsigned int *c_T, unsigned int *c_C, unsigned int *c_G, unsigned int *w_Aq, unsigned int *w_Tq, unsigned int *w_Cq, unsigned int *w_Gq, unsigned int *c_Aq, unsigned int *c_Tq, unsigned int *c_Cq, unsigned int *c_Gq, unsigned int *w_Q, unsigned int *c_Q)
{
int i, j, m;
int v, n;
float vRate[3];
// Record snp sites
for(i = 0; i < len; i++)
{
m = w_A[i]+w_T[i]+w_C[i]+w_G[i]+c_A[i]+c_T[i]+c_C[i]+c_G[i];
// No snp pt number
switch(chrSeqArray[idx][i])
{
case 'A':
// A>C=(C+WsT)/total reads
v = w_C[i] + c_C[i] + w_T[i];
n = m;
vRate[0] = (float)v/n;
// A>T=T/total reads
v = w_T[i] + c_T[i];
n = m;
vRate[1] = (float)v/n;
// A>G=WsG/(WsG+WsA)
v = w_G[i];
n = w_G[i] + w_A[i];
vRate[2] = (float)v/n;
break;
case 'T':
// T>A=A/total reads
v = w_A[i] + c_A[i];
n = m;
vRate[0] = (float)v/n;
// T>C=CrC/(CrT+CrC)
v = c_C[i];
n = c_T[i] + c_C[i];;
vRate[1] = (float)v/n;
// T>G=(G+CrA)/total reads
v = w_G[i] + c_G[i] + c_A[i];
n = m;
vRate[2] = (float)v/n;
break;
case 'C':
// C>A=A/total reads
v = w_A[i] + c_A[i];
n = m;
vRate[0] = (float)v/n;
// C>T=CrT/(CrC+CrT)
v = c_T[i];
n = c_C[i] + c_T[i];
vRate[1] = (float)v/n;
// C>G=(G+CrA)/total reads
v = w_G[i] + c_G[i] + c_A[i];
n = m;
vRate[2] = (float)v/n;
break;
case 'G':
// G>A=WsA/(WsA+WsG)
v = w_A[i];
n = w_A[i] + w_G[i];
vRate[0] = (float)v/n;
// G>T=T/total reads
v = w_T[i] + c_T[i];
n = m;
vRate[1] = (float)v/n;
// G>C=(WsT+C)/total reads
v = w_T[i] + w_C[i] + c_C[i];
n = m;
vRate[2] = (float)v/n;
break;
}
// Filtering
if(vRate[0] > vSnpRate || vRate[1] > vSnpRate || vRate[2] > vSnpRate)
{
fprintf(posFptr, "%s\t%d\t%c\t", curChr, i + 1, chrSeqArray[idx][i]);
fprintf(posFptr, "%u,%u,%u,%u\t", w_A[i], w_T[i], w_C[i], w_G[i]);
fprintf(posFptr, "%u,%u,%u,%u\t", c_A[i], c_T[i], c_C[i], c_G[i]);
fprintf(posFptr, "%u,%u,%u,%u\t", (unsigned int)((float)w_Aq[i]/w_A[i]+0.5), (unsigned int)((float)w_Tq[i]/w_T[i]+0.5), (unsigned int)((float)w_Cq[i]/w_C[i]+0.5), (unsigned int)((float)w_Gq[i]/w_G[i]+0.5));
fprintf(posFptr, "%u,%u,%u,%u\t", (unsigned int)((float)c_Aq[i]/c_A[i]+0.5), (unsigned int)((float)c_Tq[i]/c_T[i]+0.5), (unsigned int)((float)c_Cq[i]/c_C[i]+0.5), (unsigned int)((float)c_Gq[i]/c_G[i]+0.5));
fprintf(posFptr, "%u\t", (unsigned int)((float)w_Q[i]/(w_A[i]+w_T[i]+w_C[i]+w_G[i])+0.5));
fprintf(posFptr, "%u\n", (unsigned int)((float)c_Q[i]/(c_A[i]+c_T[i]+c_C[i]+c_G[i])+0.5));
}
}
}
int parseBuffer(bam_header_t *header, bam1_t *b, MapRecord* record, unsigned int mapqThr)
{
int i, num, plen, flag;
int seqPtr, seqBufPtr;
char nchar;
char *pdest;
const bam1_core_t *c = &b->core;
uint8_t *s = bam1_seq(b);
uint8_t *t = bam1_qual(b);
/////////////////////////////////
// Load record
/////////////////////////////////
// {{
// 1 - QNAME
strcpy(record->qname, bam1_qname(b));
// 2 - FLAG
record->flag = c->flag;
if((record->flag & 0x04) != 0) {
// Unmap
return 1;
}
if((record->flag & 0x10) == 0) {
record->strand = '+';
}
else {
record->strand = '-';
}
if((record->flag & 0x80) != 0) {
record->r12 = 2;
}
else {
record->r12 = 1;
}
// 3 - RNAME
strcpy(record->chrome, header->target_name[c->tid]);
// 4 - POS
record->offset = c->pos + 1;
// 5 - MAPQ
record->mapq = c->qual;
if(c->qual < mapqThr)
return 1;
// 6 - CIGAR
for (i = 0; i < c->n_cigar; ++i)
sprintf(record->cigar, "%d%c", bam1_cigar(b)[i]>>BAM_CIGAR_SHIFT, "MIDNSHP"[bam1_cigar(b)[i]&BAM_CIGAR_MASK]);
// 10 - SEQ
for(i = 0; i < c->l_qseq; ++i)
record->seqBuf[i] = bam_nt16_rev_table[bam1_seqi(s, i)];
record->seqBuf[i] = '\0';
// 11 - QUAL
if(t[0] == 0xff) {
strcpy(record->qualBuf, "*");
}
else {
for(i = 0; i < c->l_qseq; ++i)
record->qualBuf[i] = (char)(t[i] + 33);
}
// }}
/////////////////////////////////
// To upper
/////////////////////////////////
// {{
for(i = 0; i < strlen(record->seqBuf); i++)
record->seqBuf[i] = toupper(record->seqBuf[i]);
// }}
/////////////////////////////////
// Cigar analysis
/////////////////////////////////
// {{
pdest = record->cigar;
nchar = nxtChar(pdest, &plen);
seqPtr = 0;
seqBufPtr = 0;
while(nchar != 'Z') {
// Digit parsing
for(i = 0; i < plen; i++)
record->comBuf[i] = pdest[i];
record->comBuf[plen] = '\0';
num = atoi(record->comBuf);
// Sequence handling
switch(nchar) {
case 'M':
for(i = 0; i < num; i++) {
record->seq[seqPtr] = record->seqBuf[seqBufPtr];
record->qual[seqPtr] = record->qualBuf[seqBufPtr];
seqPtr++;
seqBufPtr++;
}
break;
case 'D':
for(i = 0; i < num; i++) {
record->seq[seqPtr] = 'N';
seqPtr++;
}
break;
case 'I':
seqBufPtr = seqBufPtr + num;
break;
case 'S':
for(i = 0; i < num; i++) {
record->seq[seqPtr] = 'N';
seqPtr++;
seqBufPtr++;
}
break;
default:
fprintf(stderr, "Error in cigar parsing!\n");
exit(1);
}
pdest = pdest + plen + 1;
nchar = nxtChar(pdest, &plen);
}
record->len = seqPtr;
// }}
return 0;
}
char nxtChar(char* cigar, int* len)
{
// "len" is the length of digit part
int tLen, mLen;
char mChar;
char* pdest;
mLen = 1000;
mChar = 'Z';
pdest = strstr(cigar, "M");
if(pdest != NULL) {
tLen = (int)(pdest - cigar);
if(tLen < mLen) {
mLen = tLen;
mChar = 'M';
}
}
pdest = strstr(cigar, "D");
if(pdest != NULL) {
tLen = (int)(pdest - cigar);
if(tLen < mLen) {
mLen = tLen;
mChar = 'D';
}
}
pdest = strstr(cigar, "I");
if(pdest != NULL) {
tLen = (int)(pdest - cigar);
if(tLen < mLen) {
mLen = tLen;
mChar = 'I';
}
}
pdest = strstr(cigar, "S");
if(pdest != NULL) {
tLen = (int)(pdest - cigar);
if(tLen < mLen) {
mLen = tLen;
mChar = 'S';
}
}
if(mLen < 1000)
*len = mLen;
return mChar;
}
void dispRecord(MapRecord* record)
{
int i;
for(i = 0; i < 40; i++)
fprintf(stderr, "*");
fprintf(stderr, "\n");
fprintf(stderr, "qname: %s\n", record->qname);
fprintf(stderr, "strand: %c\n", record->strand);
fprintf(stderr, "chrome: %s\n", record->chrome);
fprintf(stderr, "flag: %d\n", record->flag);
fprintf(stderr, "offset: %d\n", record->offset);
fprintf(stderr, "mapq: %d\n", record->mapq);
fprintf(stderr, "cigar: %s\n", record->cigar);
fprintf(stderr, "seq: %s\n", record->seq);
fprintf(stderr, "qual: %s\n", record->qual);
fprintf(stderr, "len: %d\n", record->len);
fprintf(stderr, "r12: %d\n", record->r12);
for(i = 0; i < 40; i++)
fprintf(stderr, "*");
fprintf(stderr, "\n");
}
int seqReverse(char* seq, int seqLen)
{
int i, j;
char tmp;
// Step 1, Reverse
for(i = 0; i < seqLen / 2; i++) {
tmp = seq[i];
seq[i] = seq[seqLen - 1 - i];
seq[seqLen - 1 - i] = tmp;
}
// Step 2, Change
for(i = 0; i < seqLen; i++) {
switch(seq[i]) {
case 'A':
seq[i] = 'T';
break;
case 'T':
seq[i] = 'A';
break;
case 'C':
seq[i] = 'G';
break;
case 'G':
seq[i] = 'C';
break;
case 'N':
seq[i] = 'N';
break;
default:
fprintf(stderr, "Sequence content error!\n");
for(j = 0; j < seqLen; j++)
fprintf(stderr, "%c", seq[j]);
fprintf(stderr, "\n");
return 1;
}
}
return 0;
}
void methProcess(char* bamFileName, char* methCgFileName, char* methChgFileName, char* methChhFileName, HashNode** hashTable, char** chrSeqArray, int* chrLen, int chrCnt, int vQualMin, int nLayerMin, unsigned int mapqThr)
{
int i, j, off, idx, len;
int m, n, cnt, iread;
int* chrDone = (int*)calloc(chrCnt, sizeof(int));
MapRecord* record = (MapRecord*)malloc(sizeof(MapRecord));
record->qname = (char*)malloc(sizeof(char) * 1000);
record->chrome = (char*)malloc(sizeof(char) * 1000);
record->cigar = (char*)malloc(sizeof(char) * 1000);
record->seq = (char*)malloc(sizeof(char) * 1000);
record->seqBuf = (char*)malloc(sizeof(char) * 1000);
record->qual = (char*)malloc(sizeof(char) * 1000);
record->qualBuf = (char*)malloc(sizeof(char) * 1000);
record->comBuf = (char*)malloc(sizeof(char) * 1000);
unsigned int *w_Mm_CG = NULL;
unsigned int *w_Mc_CG = NULL;
unsigned int *w_Mq_CG = NULL;
unsigned int *c_Mm_CG = NULL;
unsigned int *c_Mc_CG = NULL;
unsigned int *c_Mq_CG = NULL;
unsigned int *w_Mm_CHG = NULL;
unsigned int *w_Mc_CHG = NULL;
unsigned int *w_Mq_CHG = NULL;
unsigned int *c_Mm_CHG = NULL;
unsigned int *c_Mc_CHG = NULL;
unsigned int *c_Mq_CHG = NULL;
unsigned int *w_Mm_CHH = NULL;
unsigned int *w_Mc_CHH = NULL;
unsigned int *w_Mq_CHH = NULL;
unsigned int *c_Mm_CHH = NULL;
unsigned int *c_Mc_CHH = NULL;
unsigned int *c_Mq_CHH = NULL;
int rowCnt = 0;
int finCnt = 0;
char curChr[100] = "chr1234567890";
uint8_t *s, *t;
bam_header_t *header;
fprintf(stderr, "Meth process begins...\n");
bamFile in = bam_open(bamFileName, "r");
if(in == NULL) {
fprintf(stderr, "Cannot open bam file!\n");
exit(1);
}
FILE* methCgFptr = fopen(methCgFileName, "w");
if(methCgFptr == NULL) {
fprintf(stderr, "Could not open file %s!\n", methCgFileName);
exit(1);
}
fprintf(methCgFptr, "#CHROM\tPOS\tCONTEXT\tWatson-METH\tWatson-COVERAGE\tWatson-QUAL\tCrick-METH\tCrick-COVERAGE\tCrick-QUAL\n");
FILE* methChgFptr = fopen(methChgFileName, "w");
if(methChgFptr == NULL) {
fprintf(stderr, "Could not open file %s!\n", methChgFileName);
exit(1);
}
fprintf(methChgFptr, "#CHROM\tPOS\tCONTEXT\tWatson-METH\tWatson-COVERAGE\tWatson-QUAL\tCrick-METH\tCrick-COVERAGE\tCrick-QUAL\n");
FILE* methChhFptr = fopen(methChhFileName, "w");
if(methChhFptr == NULL) {
fprintf(stderr, "Could not open file %s!\n", methChhFileName);
exit(1);
}
fprintf(methChhFptr, "#CHROM\tPOS\tCONTEXT\tWatson-METH\tWatson-COVERAGE\tWatson-QUAL\tCrick-METH\tCrick-COVERAGE\tCrick-QUAL\n");
bam1_t* b = bam_init1();
if(b == NULL) {
fprintf(stderr, "Cannot init bam structure!\n");
exit(1);
}
header = bam_header_read(in);
while(bam_read1(in, b) >= 0) {
// Parse record
if(parseBuffer(header, b, record, mapqThr) == 1)
continue;
// Check chrome
if(strcmp(curChr, record->chrome) != 0) {
fprintf(stderr, "Chrome updated to %s!\n", record->chrome);
// Save old chrome statics results
if(rowCnt > 0) {
// Update
finCnt = rowCnt;
// Print
printMeth(methCgFptr, len, curChr, w_Mm_CG, w_Mc_CG, w_Mq_CG, c_Mm_CG, c_Mc_CG, c_Mq_CG, "CG", nLayerMin);
printMeth(methChgFptr, len, curChr, w_Mm_CHG, w_Mc_CHG, w_Mq_CHG, c_Mm_CHG, c_Mc_CHG, c_Mq_CHG, "CHG", nLayerMin);
printMeth(methChhFptr, len, curChr, w_Mm_CHH, w_Mc_CHH, w_Mq_CHH, c_Mm_CHH, c_Mc_CHH, c_Mq_CHH, "CHH", nLayerMin);
// Memory gathering for x_Mx_CG
free(w_Mm_CG);
free(w_Mc_CG);
free(w_Mq_CG);
free(c_Mm_CG);
free(c_Mc_CG);
free(c_Mq_CG);
// Memory gathering for x_Mx_CHG
free(w_Mm_CHG);
free(w_Mc_CHG);
free(w_Mq_CHG);
free(c_Mm_CHG);
free(c_Mc_CHG);
free(c_Mq_CHG);
// Memory gathering for x_Mx_CHH
free(w_Mm_CHH);
free(w_Mc_CHH);
free(w_Mq_CHH);
free(c_Mm_CHH);
free(c_Mc_CHH);
free(c_Mq_CHH);
}
// Update current chrome
strcpy(curChr, record->chrome);
// Get chrome length
idx = hash_table_lookup(hashTable, curChr);
if(idx == -1) {
fprintf(stderr, "%s not found in chrome name array.\n", curChr);
exit(1);
}
len = chrLen[idx];
// Check if chrome has been processed
if(chrDone[idx] != 0) {
fprintf(stderr, "%s has already been processed. The bam file is not sorted.\n", curChr);
exit(1);
}
chrDone[idx] = 1;
// Memory alloction for x_Mx_CG
if(!(w_Mm_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mc_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mq_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mq_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mm_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mc_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mq_CG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mq_CG of %s.\n", curChr);
exit(1);
}
// Memory alloction for x_Mx_CHG
if(!(w_Mm_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mc_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mq_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mq_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mm_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mc_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mq_CHG = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mq_CG of %s.\n", curChr);
exit(1);
}
// Memory alloction for x_Mx_CG
if(!(w_Mm_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mc_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(w_Mq_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Mq_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mm_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mm_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mc_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mc_CG of %s.\n", curChr);
exit(1);
}
if(!(c_Mq_CHH = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Mq_CG of %s.\n", curChr);
exit(1);
}
}
/////////////////////////////////////////////////
// Filter Step 1: Base quality
/////////////////////////////////////////////////
for(i = 0; i < record->len; i++) {
if(record->seq[i] != 'N' && (unsigned short)(record->qual[i] - 33) < vQualMin)
record->seq[i] = 'N';
}
/////////////////////////////////////////////////
// Additional Step 1: Check and record CG methy
/////////////////////////////////////////////////
off = record->offset - 1;
if((record->r12 == 1 && record->strand == '+') || (record->r12 == 2 && record->strand == '-')) {
// Watson chain
for(i = 0; i < record->len; i++) {
if(off >= 0 && off + 1 < chrLen[idx]) {
if(record->seq[i] == 'C') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] == 'G') {
w_Mm_CG[off]++;
w_Mc_CG[off]++;
w_Mq_CG[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'T') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] == 'G') {
w_Mc_CG[off]++;
w_Mq_CG[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
else {
// Crick chain
for(i = 0; i < record->len; i++) {
if(off - 1 >= 0 && off < chrLen[idx]) {
if(record->seq[i] == 'G') {
//fprintf(stderr, "read[%d] = G, ref[%d-%d] = %c%c.\n", i+1, off-1, off, chrSeqArray[idx][off-1], chrSeqArray[idx][off]);
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] == 'C') {
c_Mm_CG[off]++;
c_Mc_CG[off]++;
c_Mq_CG[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'A') {
//fprintf(stderr, "read[%d] = A, ref[%d-%d] = %c%c.\n", i+1, off-1, off, chrSeqArray[idx][off-1], chrSeqArray[idx][off]);
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] == 'C') {
c_Mc_CG[off]++;
c_Mq_CG[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
/////////////////////////////////////////////////
// Additional Step 2: Check and record CHG methy
/////////////////////////////////////////////////
off = record->offset - 1;
if((record->r12 == 1 && record->strand == '+') || (record->r12 == 2 && record->strand == '-')) {
// Watson chain
for(i = 0; i < record->len; i++) {
if(off >= 0 && off + 2 < chrLen[idx]) {
if(record->seq[i] == 'C') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] != 'G' && chrSeqArray[idx][off+2] == 'G') {
w_Mm_CHG[off]++;
w_Mc_CHG[off]++;
w_Mq_CHG[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'T') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] != 'G' && chrSeqArray[idx][off+2] == 'G') {
w_Mc_CHG[off]++;
w_Mq_CHG[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
else {
// Crick chain
for(i = 0; i < record->len; i++) {
if(off - 2 >= 0 && off < chrLen[idx]) {
if(record->seq[i] == 'G') {
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] != 'C' && chrSeqArray[idx][off-2] == 'C') {
c_Mm_CHG[off]++;
c_Mc_CHG[off]++;
c_Mq_CHG[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'A') {
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] != 'C' && chrSeqArray[idx][off-2] == 'C') {
c_Mc_CHG[off]++;
c_Mq_CHG[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
/////////////////////////////////////////////////
// Additional Step 3: Check and record CHH methy
/////////////////////////////////////////////////
off = record->offset - 1;
if((record->r12 == 1 && record->strand == '+') || (record->r12 == 2 && record->strand == '-')) {
// Watson chain
for(i = 0; i < record->len; i++) {
/*
if(off == 257561 && strcmp(curChr, "chr1") == 0) {
fprintf(stderr, "\n\n");
for(j = 0; j < 10; j++)
fprintf(stderr, "%c", chrSeqArray[idx][off+j]);
fprintf(stderr, "\n\n");
fprintf(stderr, "%d, %c, %c, %d\n", record->r12, record->strand, record->seq[i], chrLen[idx]);
exit(1);
}
*/
if(off >= 0 && off + 2 < chrLen[idx]) {
if(record->seq[i] == 'C') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] != 'G' && chrSeqArray[idx][off+2] != 'G') {
w_Mm_CHH[off]++;
w_Mc_CHH[off]++;
w_Mq_CHH[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'T') {
if(chrSeqArray[idx][off] == 'C' && chrSeqArray[idx][off+1] != 'G' && chrSeqArray[idx][off+2] != 'G') {
w_Mc_CHH[off]++;
w_Mq_CHH[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
else {
// Crick chain
for(i = 0; i < record->len; i++) {
if(off - 2 >= 0 && off < chrLen[idx]) {
if(record->seq[i] == 'G') {
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] != 'C' && chrSeqArray[idx][off-2] != 'C') {
c_Mm_CHH[off]++;
c_Mc_CHH[off]++;
c_Mq_CHH[off] += record->qual[i] - 33;
}
}
else if(record->seq[i] == 'A') {
if(chrSeqArray[idx][off] == 'G' && chrSeqArray[idx][off-1] != 'C' && chrSeqArray[idx][off-2] != 'C') {
c_Mc_CHH[off]++;
c_Mq_CHH[off] += record->qual[i] - 33;
}
}
}
off++;
}
}
// Update row counter
rowCnt++;
if(rowCnt % 500000 == 0)
fprintf(stderr, "%d records have been dealed.\n", rowCnt);
}
// Last batch
if(finCnt < rowCnt) {
// Print
printMeth(methCgFptr, len, curChr, w_Mm_CG, w_Mc_CG, w_Mq_CG, c_Mm_CG, c_Mc_CG, c_Mq_CG, "CG", nLayerMin);
printMeth(methChgFptr, len, curChr, w_Mm_CHG, w_Mc_CHG, w_Mq_CHG, c_Mm_CHG, c_Mc_CHG, c_Mq_CHG, "CHG", nLayerMin);
printMeth(methChhFptr, len, curChr, w_Mm_CHH, w_Mc_CHH, w_Mq_CHH, c_Mm_CHH, c_Mc_CHH, c_Mq_CHH, "CHH", nLayerMin);
// Memory gathering for x_Mx_CG
free(w_Mm_CG);
free(w_Mc_CG);
free(w_Mq_CG);
free(c_Mm_CG);
free(c_Mc_CG);
free(c_Mq_CG);
// Memory gathering for x_Mx_CHG
free(w_Mm_CHG);
free(w_Mc_CHG);
free(w_Mq_CHG);
free(c_Mm_CHG);
free(c_Mc_CHG);
free(c_Mq_CHG);
// Memory gathering for x_Mx_CHH
free(w_Mm_CHH);
free(w_Mc_CHH);
free(w_Mq_CHH);
free(c_Mm_CHH);
free(c_Mc_CHH);
free(c_Mq_CHH);
}
bam_header_destroy(header);
bam_close(in);
bam_destroy1(b);
fclose(methCgFptr);
fclose(methChgFptr);
fclose(methChhFptr);
fprintf(stderr, "Meth process ends...\n");
}
void snpProcess(char* bamFileName, char* snpFileName, HashNode** hashTable, char** chrSeqArray, int* chrLen, int chrCnt, int vQualMin, int nLayerMax, float vSnpRate, float vSnpPerBase, unsigned int mapqThr)
{
int i, j, off, idx, len;
int m, n, cnt, iread;
int* chrDone = (int*)calloc(chrCnt, sizeof(int));
MapRecord* record = (MapRecord*)malloc(sizeof(MapRecord));
record->qname = (char*)malloc(sizeof(char) * 1000);
record->chrome = (char*)malloc(sizeof(char) * 1000);
record->cigar = (char*)malloc(sizeof(char) * 1000);
record->seq = (char*)malloc(sizeof(char) * 1000);
record->seqBuf = (char*)malloc(sizeof(char) * 1000);
record->qual = (char*)malloc(sizeof(char) * 1000);
record->qualBuf = (char*)malloc(sizeof(char) * 1000);
record->comBuf = (char*)malloc(sizeof(char) * 1000);
unsigned int *w_A = NULL;
unsigned int *w_T = NULL;
unsigned int *w_C = NULL;
unsigned int *w_G = NULL;
unsigned int *c_A = NULL;
unsigned int *c_T = NULL;
unsigned int *c_C = NULL;
unsigned int *c_G = NULL;
unsigned int *w_Aq = NULL;
unsigned int *w_Tq = NULL;
unsigned int *w_Cq = NULL;
unsigned int *w_Gq = NULL;
unsigned int *c_Aq = NULL;
unsigned int *c_Tq = NULL;
unsigned int *c_Cq = NULL;
unsigned int *c_Gq = NULL;
unsigned int *w_Q = NULL;
unsigned int *c_Q = NULL;
int rowCnt = 0;
int finCnt = 0;
char curChr[100] = "chr1234567890";
uint8_t *s, *t;
bam_header_t *header;
fprintf(stderr, "SNP process begins...\n");
bamFile in = bam_open(bamFileName, "r");
if(in == NULL) {
fprintf(stderr, "Cannot open bam file!\n");
exit(1);
}
FILE* snpFptr = fopen(snpFileName, "w");
if(snpFptr == NULL) {
fprintf(stderr, "Could not open file %s!\n", snpFileName);
exit(1);
}
fprintf(snpFptr, "#CHROM\tPOS\tID\tREF\tALT\tQUAL\tFILTER\tGENOTYPE\tFREQUENCY\tNumber_of_watson[A,T,C,G]\tNumber_of_crick[A,T,C,G]\tMean_Quality_of_Watson[A,T,C,G]\tMean_Quality_of_Crick[A,T,C,G]\n");
bam1_t* b = bam_init1();
if(b == NULL) {
fprintf(stderr, "Cannot init bam structure!\n");
exit(1);
}
header = bam_header_read(in);
while(bam_read1(in, b) >= 0) {
// Parse record
if(parseBuffer(header, b, record, mapqThr) == 1)
continue;
// Check chrome
if(strcmp(curChr, record->chrome) != 0) {
// Save old chrome statics results
if(rowCnt > 0) {
// Update
finCnt = rowCnt;
// Print
printSnp(snpFptr, chrSeqArray, idx, len, vSnpRate, curChr, w_A, w_T, w_C, w_G, c_A, c_T, c_C, c_G, w_Aq, w_Tq, w_Cq, w_Gq, c_Aq, c_Tq, c_Cq, c_Gq, w_Q, c_Q);
// Memory gathering for x_X
free(w_A);
free(w_T);
free(w_C);
free(w_G);
free(c_A);
free(c_T);
free(c_C);
free(c_G);
// Memory gathering for x_Xq
free(w_Aq);
free(w_Tq);
free(w_Cq);
free(w_Gq);
free(c_Aq);
free(c_Tq);
free(c_Cq);
free(c_Gq);
// Memory gathering for x_Q
free(w_Q);
free(c_Q);
}
// Update current chrome
strcpy(curChr, record->chrome);
// Get chrome length
idx = hash_table_lookup(hashTable, curChr);
if(idx == -1) {
fprintf(stderr, "%s not found in chrome name array.\n", curChr);
exit(1);
}
len = chrLen[idx];
// Check if chrome has been processed
if(chrDone[idx] != 0) {
fprintf(stderr, "%s has already been processed. The bam file is not sorted.\n", curChr);
exit(1);
}
chrDone[idx] = 1;
// Memory alloction for x_X
if(!(w_A = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_A of %s.\n", curChr);
exit(1);
}
if(!(w_T = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_T of %s.\n", curChr);
exit(1);
}
if(!(w_C = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_C of %s.\n", curChr);
exit(1);
}
if(!(w_G = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_G of %s.\n", curChr);
exit(1);
}
if(!(c_A = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_A of %s.\n", curChr);
exit(1);
}
if(!(c_T = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_T of %s.\n", curChr);
exit(1);
}
if(!(c_C = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_C of %s.\n", curChr);
exit(1);
}
if(!(c_G = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_G of %s.\n", curChr);
exit(1);
}
// Memory alloction for x_Xq
if(!(w_Aq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Aq of %s.\n", curChr);
exit(1);
}
if(!(w_Tq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Tq of %s.\n", curChr);
exit(1);
}
if(!(w_Cq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Cq of %s.\n", curChr);
exit(1);
}
if(!(w_Gq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Gq of %s.\n", curChr);
exit(1);
}
if(!(c_Aq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Aq of %s.\n", curChr);
exit(1);
}
if(!(c_Tq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Tq of %s.\n", curChr);
exit(1);
}
if(!(c_Cq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Cq of %s.\n", curChr);
exit(1);
}
if(!(c_Gq = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Gq of %s.\n", curChr);
exit(1);
}
// Memory alloction for x_Q
if(!(w_Q = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for w_Q of %s.\n", curChr);
exit(1);
}
if(!(c_Q = (unsigned int*)calloc(len, sizeof(unsigned int)))) {
fprintf(stderr, "Not enough memory for c_Q of %s.\n", curChr);
exit(1);
}
}
/////////////////////////////////////////////////
// Filter Step 1: Base quality
/////////////////////////////////////////////////