From d4f14048de7b2b1b75160d45196dbc9c82828be5 Mon Sep 17 00:00:00 2001 From: David Jones Date: Wed, 24 Aug 2016 11:20:37 +0100 Subject: [PATCH 1/8] Added bam2bw functionality. --- c/Makefile | 15 +- c/bam2bedgraph.c | 2 +- c/bam2bw.c | 518 +++++++++++++++++++++++++++++++++++++++++++++++ setup.sh | 1 + 4 files changed, 531 insertions(+), 5 deletions(-) create mode 100644 c/bam2bw.c diff --git a/c/Makefile b/c/Makefile index 8d5c8b4..818c4b0 100644 --- a/c/Makefile +++ b/c/Makefile @@ -67,6 +67,7 @@ BAM_STATS_TARGET=../bin/bam_stats CAT_TARGET=../bin/bwcat SQ_TARGET=../bin/reheadSQ BAM2BG_TARGET=../bin/bam2bedgraph +BAM2BW_TARGET=../bin/bam2bw # # The following part of the makefile is generic; it can be used to @@ -78,8 +79,8 @@ BAM2BG_TARGET=../bin/bam2bedgraph .NOTPARALLEL: test -all: clean pre make_htslib_tmp $(BAM_STATS_TARGET) $(BAM2BG_TARGET) test remove_htslib_tmp $(CAT_TARGET) $(SQ_TARGET) - @echo bam_stats, reheadSQ and bwcat compiled. +all: clean pre make_htslib_tmp $(BAM_STATS_TARGET) $(BAM2BG_TARGET) $(BAM2BW_TARGET) test remove_htslib_tmp $(CAT_TARGET) $(SQ_TARGET) + @echo bam_stats, reheadSQ, bwcat and bam2bw compiled. $(BAM_STATS_TARGET): $(OBJS) $(CC) $(CFLAGS) $(INCLUDES) -o $(BAM_STATS_TARGET) $(OBJS) $(LFLAGS) $(LIBS) ./bam_stats.c @@ -87,6 +88,9 @@ $(BAM_STATS_TARGET): $(OBJS) $(CAT_TARGET): $(CC) $(CAT_INCLUDES) $(CFLAGS) ./bwcat.c $(CAT_LFLAGS) -lBigWig -lz -lcurl -lm -lgnutls -ltasn1 -lhogweed -lnettle -lgmp -lp11-kit -o $(CAT_TARGET) +$(BAM2BW_TARGET): + $(CC) $(CAT_INCLUDES) $(INCLUDES) $(CFLAGS) ./bam2bw.c $(CAT_LFLAGS) -lBigWig -lz -lcurl -lm -lgnutls -ltasn1 -lhogweed -lnettle -lgmp -lp11-kit $(LFLAGS) $(LIBS) -o $(BAM2BW_TARGET) + $(BAM2BG_TARGET): $(OBJS) $(CC) $(CFLAGS) $(INCLUDES) -o $(BAM2BG_TARGET) $(OBJS) $(LFLAGS) $(LIBS) ./bam2bedgraph.c @@ -104,6 +108,9 @@ test: $(TESTS) coverage: CFLAGS += --coverage coverage: test +make_bam2bw: clean pre make_htslib_tmp $(BAM2BW_TARGET) remove_htslib_tmp + @echo $(BAM2BW_TARGET) done + make_bigwigcat: $(CAT_TARGET) @echo $(CAT_TARGET) done @@ -118,7 +125,7 @@ remove_htslib_tmp: copyscript: cp ./scripts/* ./bin/ - chmod a+x $(BAM_STATS_TARGET) $(CAT_TARGET) $(SQ_TARGET) + chmod a+x $(BAM_STATS_TARGET) $(CAT_TARGET) $(SQ_TARGET) $(BAM2BW_TARGET) $(BAM2BG_TARGET) valgrind: VALGRIND="valgrind --log-file=/tmp/valgrind-%p.log" $(MAKE) @@ -133,7 +140,7 @@ valgrind: clean: @echo clean - $(RM) ./*.o *~ $(BAM_STATS_TARGET) $(SQ_TARGET) $(CAT_TARGET) ./tests/tests_log $(TESTS) ./*.gcda ./*.gcov ./*.gcno *.gcda *.gcov *.gcno ./tests/*.gcda ./tests/*.gcov ./tests/*.gcno + $(RM) ./*.o *~ $(BAM_STATS_TARGET) $(SQ_TARGET) $(CAT_TARGET) $(BAM2BW_TARGET) $(BAM2BG_TARGET) ./tests/tests_log $(TESTS) ./*.gcda ./*.gcov ./*.gcno *.gcda *.gcov *.gcno ./tests/*.gcda ./tests/*.gcov ./tests/*.gcno -rm -rf $(HTSTMP) depend: $(SRCS) diff --git a/c/bam2bedgraph.c b/c/bam2bedgraph.c index a78cc05..7b97790 100644 --- a/c/bam2bedgraph.c +++ b/c/bam2bedgraph.c @@ -208,7 +208,7 @@ int main(int argc, char *argv[]){ } //Check we've written everything... - int ret, n_plp, tid, pos; + int n_plp, tid, pos; const bam_pileup1_t *plp; while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ pileup_func(tid, pos, n_plp, plp, &tmp); diff --git a/c/bam2bw.c b/c/bam2bw.c new file mode 100644 index 0000000..3918963 --- /dev/null +++ b/c/bam2bw.c @@ -0,0 +1,518 @@ +#include +#include +#include +#include +#include +#include +#include "bigWig.h" +#include "htslib/hts.h" +#include "htslib/sam.h" +#include "dbg.h" + +char *out_file = "output.bam.bw"; +char *input_file = NULL; +char *region_store = NULL; +char **our_region_list = NULL; +uint32_t region_list_count = 0; +int is_regions_file = 0; +uint8_t base_bit = 0; +int filter = 0; +char base = 0; +uint32_t single = 1; + +typedef struct { + uint32_t ltid; + int lstart,lcoverage,lpos,beg,end; + htsFile *in; + hts_idx_t *idx; + bam_hdr_t *head; + bigWigFile_t *out; +} tmpstruct_t; + +void print_version (int exit_code){ + printf ("%s\n",VERSION); + exit(exit_code); +} + +int line_count (char *file_path){ + FILE *f = fopen(file_path,"r"); + int line_count = 0; + check(f != NULL, "Error opening file '%s' to count lines.",file_path); + char rd[512]; + while(fgets(rd, 512, f) != NULL){ + line_count++; + } + fclose(f); + return line_count; +error: + if(f) fclose(f); + return -1; +} + +int check_exist(char *fname){ + FILE *fp; + if((fp = fopen(fname,"r"))){ + fclose(fp); + return 1; + } + return 0; +} + +void print_usage (int exit_code){ + printf("Usage: bam2bw -i input.[b|cr]am -o output.bw\n\n"); + printf("-i --input [file] Path to the input [b|cr]am file.\n"); + printf("-F --filter [int] SAM flags to filter. [default: %d]\n",filter); + printf("-o --outfile [file] Path to the output .bw file produced. [default:'%s']\n\n",out_file); + printf("Optional: \n"); + printf("-b --base [char] To produce a single base coverage bigwig file [ACTG]. Must be used in conjunction with -r|--region\n"); + printf("-r --region [file] A samtools style region (contig:start-stop) or a bed file of regions over which to produce the bigwig file\n\n"); + printf ("Other:\n"); + printf("-h --help Display this usage information.\n"); + printf("-v --version Prints the version number.\n\n"); + + exit(exit_code); +} + +int check_region_string(char *region){ + if(check_exist(region)==1){ + is_regions_file = 1; + return 1; + }else{ + //Parse this as a region + int beg = 0; + int end = 0; + const char *res = hts_parse_reg(region, &beg, &end); + if(res) return 1; + } + return -1; +} + +int get_int_length(int input){ + return (input == 0 ? 1 : (int)(log10(input)+1)); +} + +int get_no_of_SQ_lines(char *bam_loc){ + htsFile *bam = NULL; + char *line = NULL; + bam_hdr_t *header = NULL; + bam = hts_open(bam_loc, "r"); + check(bam != 0,"Bam file %s failed to open to read header.",bam_loc); + header = sam_hdr_read(bam); + char *head_txt = header->text; + line = strtok(head_txt,"\n"); + int count = 0; + while(line != NULL){ + //Check for a read group line + if(strncmp(line,"@SQ",3)==0){ + count++; + }//End of if this is an SQ line + line = strtok(NULL,"\n"); + } + free(line); + hts_close(bam); + bam_hdr_destroy(header); + return count; +error: + if(line) free(line); + if(bam) hts_close(bam); + if(header) bam_hdr_destroy(header); + return -1; +} + +int parse_SQ_line(char *line, char *name, int *length){ + char *tag = NULL; + char *tmp = NULL; + char *ptr = NULL; + int nom = 0; + int len = 0; + tag = strtok_r(line,"\t",&ptr); + while(tag != NULL){ + int chk=0; + tmp = malloc(sizeof(char) * 512); + check_mem(tmp); + chk = sscanf(tag,"SN:%[^\t\n]",tmp); + if(chk>0){ + strcpy(name,tmp); + tag = strtok_r(NULL,"\t",&ptr); + nom = 1; + continue; + } + chk = sscanf(tag,"LN:%d",length); + if(chk>0){ + len = 1; + tag = strtok_r(NULL,"\t",&ptr); + continue; + } + tag = strtok_r(NULL,"\t",&ptr); + } + if(tmp) free(tmp); + if(tag) free(tag); + + if(nom && len) { + return 1; + }else{ + return -1; + } +error: + if(tmp) free(tmp); + if(tag) free(tag); + return -1; +} + +int build_chromList_from_bam(chromList_t *chromList ,char *bam_loc){ + htsFile *bam = NULL; + char *line = NULL; + char *tmp = NULL; + bam_hdr_t *header = NULL; + char *ptr; + bam = hts_open(bam_loc, "r"); + check(bam != 0,"Bam file %s failed to open to read header.",bam_loc); + header = sam_hdr_read(bam); + char *head_txt = header->text; + line = strtok(head_txt,"\n"); + int i = 0; + while(line != NULL){ + //Check for a read group line + if(strncmp(line,"@SQ",3)==0){ + char *tag = strtok_r(line,"\t",&ptr); + while(tag != NULL){ + int chk=0; + tmp = malloc(sizeof(char) * 512); + check_mem(tmp); + chk = sscanf(tag,"SN:%[^\t\n]",tmp); + if(chk>0){ + chromList->chrom[i] = malloc(sizeof(char) * (strlen(tmp)+1)); + check_mem(chromList->chrom[i]); + strcpy(chromList->chrom[i],tmp); + tag = strtok_r(NULL,"\t",&ptr); + free(tmp); + continue; + } + free(tmp); + uint32_t tmpint = 0; + chk = sscanf(tag,"LN:%" SCNu32 "",&tmpint); + if(chk>0){ + chromList->len[i] = tmpint; + tag = strtok_r(NULL,"\t",&ptr); + continue; + } + tag = strtok_r(NULL,"\t",&ptr); + }//End of SQ line tags + i++; + }//End of if this is an SQ line + line = strtok(NULL,"\n"); + } + free(line); + bam_hdr_destroy(header); + hts_close(bam); + return 1; +error: + if(line) free(line); + if(bam) hts_close(bam); + if(header) bam_hdr_destroy(header); + if(tmp) free (tmp); + return -1; +} + +void setup_options(int argc, char *argv[]){ + const struct option long_opts[] = + { + {"input", required_argument, 0, 'i'}, + {"filter", required_argument, 0, 'F'}, + {"outfile",required_argument, 0, 'o'}, + {"base",required_argument, 0, 'b'}, + {"region",required_argument, 0, 'r'}, + {"help", no_argument, 0, 'h'}, + {"version", no_argument, 0, 'v'}, + { NULL, 0, NULL, 0} + + }; //End of declaring opts + + int index = 0; + int iarg = 0; + + //Iterate through options + while((iarg = getopt_long(argc, argv, "F:i:o:b:r:hv",long_opts, &index)) != -1){ + switch(iarg){ + case 'F': + if(sscanf(optarg, "%i", &filter) != 1){ + fprintf(stderr,"Error parsing -F|--filter argument '%s'. Should be an integer > 0",optarg); + print_usage(1); + } + break; + case 'i': + input_file = optarg; + if(check_exist(input_file) != 1){ + fprintf(stderr,"Input bam file %s does not appear to exist.\n",input_file); + print_usage(1); + } + break; + case 'o': + out_file = optarg; + break; + case 'h': + print_usage (0); + break; + case 'v': + print_version (0); + break; + case 'r': + region_store = optarg; + //First check for a region format + int res = check_region_string(region_store); + if(res<0){ + fprintf(stderr,"Region %s is not in correct format or not an existing bed file.\n",region_store); + print_usage(1); + } + break; + case 'b': + if(sscanf(optarg, "%c", &base) != 1){ + fprintf(stderr,"Error parsing -b|--base argument '%s'. Should be a single character > 0",optarg); + print_usage(1); + } + if(base != 'A' && base != 'C' && base != 'G' && base != 'T'){ + fprintf(stderr,"Single base %c provided must match [ACGT].\n",base); + print_usage(1); + }else{ + switch (base){ + case 'A': + base_bit = 1; + break; + case 'C': + base_bit = 2; + break; + case 'G': + base_bit = 4; + break; + case 'T': + base_bit = 8; + break; + } + } + break; + case '?': + print_usage (1); + break; + default: + print_usage (0); + }; // End of args switch statement + + }//End of iteration through options + + if(input_file==NULL){ + fprintf(stderr,"Required option -i|--input-bam not defined.\n"); + print_usage(1); + } + + if(base_bit != 0 && region_store==NULL){ + fprintf(stderr,"Option -r|--region must be used with the -b|--base option.\n"); + print_usage(1); + } + + return; +} + +// callback for bam_plbuf_init() +static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data) +{ + tmpstruct_t *tmp = (tmpstruct_t*)data; + int pos = (int)position; + int coverage = n; + int i; + for (i=0;i0 && bam_seqi(bam_get_seq(pl[i].b), pl[i].qpos)!=base_bit)) coverage--; + if (tmp->ltid != tid || tmp->lcoverage != coverage || pos > tmp->lpos+1) { + if (tmp->lpos > 0 && tmp->lcoverage > 0){ + uint32_t start = tmp->lstart; + uint32_t stop = tmp->lpos+1; + float cvg = tmp->lcoverage; + bwAddIntervals(tmp->out,&tmp->head->target_name[tmp->ltid],&start,&stop,&cvg,single); + } + tmp->ltid = tid; + tmp->lstart = pos; + tmp->lcoverage = coverage; + } + tmp->lpos = pos; + return 0; +} + +int main(int argc, char *argv[]){ + setup_options(argc, argv); + tmpstruct_t tmp; + int no_of_regions = 0; + int sq_lines = get_no_of_SQ_lines(input_file); + FILE *fp_bed = NULL; + if(region_store){ + if(is_regions_file==1){ + //If we have a bedfile + no_of_regions = line_count(region_store); + check(no_of_regions>0,"Error counting entries in region file %s\n",region_store); + our_region_list = calloc(no_of_regions, sizeof(char *)); + check_mem(region_list); + int i=0; + fp_bed = fopen(region_store,"r"); + char line[512]; + while(fgets(line,512,fp_bed)){ + char *contig = malloc(sizeof(char)*256); + check_mem(contig); + int beg,end; + int chk = sscanf(line,"%s\t%d\t%d",contig,&beg,&end); + check(chk==3,"Error reading line '%s' from regions bed file.",line); + char *region = malloc(sizeof(char) * (strlen(contig)+get_int_length(beg)+get_int_length(end))+3); + check_mem(region); + chk = sprintf(region,"%s:%d-%d",contig,beg+1,end); //+1 to beginning as this is bed + check(chk==((strlen(contig)+get_int_length(beg)+get_int_length(end))+2),"Error creating region line from bed entry '%s'.",line); + our_region_list[i] = region; + i++; + } + fclose(fp_bed); + }else{ + //If we have a single region.... + no_of_regions = 1; + our_region_list = calloc(no_of_regions, sizeof(char *)); + check_mem(region_list); + our_region_list[0] = region_store; + } + }else{ + //If not... + //Build a list of regions from the header of the bamfile + no_of_regions = sq_lines; + our_region_list = calloc(no_of_regions, sizeof(char *)); + htsFile *bam = NULL; + char *line = NULL; + bam = hts_open(input_file, "r"); + check(bam != 0,"Bam file %s failed to open to read header.",input_file); + bam_hdr_t *header = sam_hdr_read(bam); + char *head_txt = header->text; + line = strtok(head_txt,"\n"); + int i=0; + while(line != NULL){ + //Check for a read group line + if(strncmp(line,"@SQ",3)==0){ + char *contig = malloc(sizeof(char) * 256); + int end,chk; + chk = parse_SQ_line(line,contig,&end); + check(chk==1,"Error parsing SQ line %s.\n",line); + char *region = malloc(sizeof(char) * (strlen(contig)+get_int_length(end))+4); + check_mem(region); + chk = sprintf(region,"%s:%d-%d",contig,1,end); + check(chk==((strlen(contig)+get_int_length(end))+3),"Error creating region line from bed entry '%s'.",line); + our_region_list[i] = region; + i++; + }//End of if this is an SQ line + line = strtok(NULL,"\n"); + } + bam_hdr_destroy(header); + free(line); + hts_close(bam); + } + + check(no_of_regions>0,"Error evaluating regions to parse. None found."); + + //Now create a bigwigfile + //Open file as a bw file + chromList_t *chromList = NULL; + bigWigFile_t *bw_out = NULL; + bam_plp_t buf = NULL; + bam1_t *b = NULL; + hts_itr_t *iter = NULL; + //Generate the list of chromosomes using the bam header. + chromList = calloc(1, sizeof(chromList_t)); + check_mem(chromList); + chromList->nKeys = sq_lines; + chromList->chrom = calloc(sq_lines,sizeof(char *)); + check_mem(chromList->chrom); + chromList->len = calloc(sq_lines,sizeof(uint32_t)); + check_mem(chromList->len); + //Iterate through the header of the bam file and get all contigs. + int res = build_chromList_from_bam(chromList,input_file); + check(res==1,"Error building chromList from bam header."); + + //Open output file + bw_out = bwOpen(out_file, NULL, "w"); + check(bw_out!=NULL, "Error opening %s for writing.",out_file); + //Initialise bw + res = bwInit(1<<17); + check(res==0,"Received an error in bwInit"); + //10 zoom levels + res = bwCreateHdr(bw_out, 10); + check(res==0,"Received error %d in bwCreateHeader",res); + //Create the chromosome lists + bw_out->cl = bwCreateChromList(chromList->chrom, chromList->len, chromList->nKeys); + check(bw_out->cl != NULL, "Error generating chromlist as bw."); + res = bwWriteHdr(bw_out); + check(res==0,"Error %d writing bw header.",res); + + tmp.beg = 0; tmp.end = 0x7fffffff; + tmp.lstart = 0; + tmp.lcoverage = 0; + tmp.ltid = 0; + tmp.lpos = 0; + tmp.out = bw_out; + + //Open input file + tmp.in = hts_open(input_file, "r"); + check(tmp.in!=0,"Failed to open [CR|B]AM file '%s'.", input_file); + tmp.head = sam_hdr_read(tmp.in); + //Now we generate the bw info + int i=0; + for(i=0;i= 0) { + if((b->core.flag & filter)>0) continue; //Skip if this is a filtered read + int ret, n_plp, tid, pos; + const bam_pileup1_t *plp; + ret = bam_plp_push(buf, b); + if (ret < 0) break; + while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ + pileup_func(tid, pos, n_plp, plp, &tmp); + } + } + bam_plp_push(buf,0); + + //Check we've written everything... + int n_plp, tid, pos; + const bam_pileup1_t *plp; + while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ + pileup_func(tid, pos, n_plp, plp, &tmp); + } + uint32_t start = tmp.lstart; + uint32_t stop = tmp.lpos+1; + float cvg = tmp.lcoverage; + bwAddIntervals(tmp.out,&tmp.head->target_name[tmp.ltid],&start,&stop,&cvg,single); + + bam_plp_destroy(buf); + bam_destroy1(b); + sam_itr_destroy(iter); + } + + bwClose(bw_out); + bwCleanup(); + hts_close(tmp.in); + int clean=0; + for(clean=0; cleanchrom){ + free(chromList->chrom[clean]); + } + } + free(chromList->len); + free(chromList); + hts_idx_destroy(tmp.idx); + bam_hdr_destroy(tmp.head); + free(our_region_list); + return 0; + +error: + if(our_region_list) free(our_region_list); + if(fp_bed) fclose(fp_bed); + return -1; +} diff --git a/setup.sh b/setup.sh index 9b1f85a..7d7ec0d 100755 --- a/setup.sh +++ b/setup.sh @@ -319,6 +319,7 @@ else cp bin/bwcat $INST_PATH/bin/. cp bin/reheadSQ $INST_PATH/bin/. cp bin/bam2bedgraph $INST_PATH/bin/. + cp bin/bam2bw $INST_PATH/bin/. touch $SETUP_DIR/bam_stats.success make -C c clean fi From 3caeed5b0754ae9a9b1bde64edebe8e222b779aa Mon Sep 17 00:00:00 2001 From: David Jones Date: Thu, 25 Aug 2016 11:38:41 +0100 Subject: [PATCH 2/8] bam2bw - generates a coverage bw file, or can be used to produce per base proportion coverage bw over a region --- c/bam2bw.c | 224 +++++++++++++++++++++++++++++++++++++++-------------- 1 file changed, 165 insertions(+), 59 deletions(-) diff --git a/c/bam2bw.c b/c/bam2bw.c index 3918963..719bf67 100644 --- a/c/bam2bw.c +++ b/c/bam2bw.c @@ -13,10 +13,11 @@ char *out_file = "output.bam.bw"; char *input_file = NULL; char *region_store = NULL; char **our_region_list = NULL; +char *reference = NULL; uint32_t region_list_count = 0; int is_regions_file = 0; -uint8_t base_bit = 0; -int filter = 0; +uint8_t is_base = 0; +int filter = 4; char base = 0; uint32_t single = 1; @@ -27,8 +28,20 @@ typedef struct { hts_idx_t *idx; bam_hdr_t *head; bigWigFile_t *out; + } tmpstruct_t; +typedef struct { + uint32_t ltid; + int lstart,lcoverage,lpos,beg,end; + float lbaseprop; + htsFile *in; + hts_idx_t *idx; + bam_hdr_t *head; + bigWigFile_t *out; + uint8_t base_bit; +} tmpstructbase_t; + void print_version (int exit_code){ printf ("%s\n",VERSION); exit(exit_code); @@ -59,13 +72,15 @@ int check_exist(char *fname){ } void print_usage (int exit_code){ - printf("Usage: bam2bw -i input.[b|cr]am -o output.bw\n\n"); + printf("Usage: bam2bw -i input.[b|cr]am -o output.bw\n"); + printf("bam2bw can be used to generate a bw file of coverage from a [cr|b]am file or a per base proportion bigwig file over a specified region.\n\n"); printf("-i --input [file] Path to the input [b|cr]am file.\n"); printf("-F --filter [int] SAM flags to filter. [default: %d]\n",filter); - printf("-o --outfile [file] Path to the output .bw file produced. [default:'%s']\n\n",out_file); + printf("-o --outfile [file] Path to the output .bw file produced. Per base results wiillbe output as four bw files [ACGT].outputname.bw [default:'%s']\n\n",out_file); printf("Optional: \n"); - printf("-b --base [char] To produce a single base coverage bigwig file [ACTG]. Must be used in conjunction with -r|--region\n"); - printf("-r --region [file] A samtools style region (contig:start-stop) or a bed file of regions over which to produce the bigwig file\n\n"); + printf("-b --perbase To produce per base coverage bigwig files [ACTG]. Must be used in conjunction with -r|--region\n"); + printf("-c --region [file] A samtools style region (contig:start-stop) or a bed file of regions over which to produce the bigwig file\n"); + printf("-r --reference [file] Path to reference genome.fa file (required for cram if ref_path cannot be resolved)\n\n"); printf ("Other:\n"); printf("-h --help Display this usage information.\n"); printf("-v --version Prints the version number.\n\n"); @@ -220,8 +235,9 @@ void setup_options(int argc, char *argv[]){ {"input", required_argument, 0, 'i'}, {"filter", required_argument, 0, 'F'}, {"outfile",required_argument, 0, 'o'}, - {"base",required_argument, 0, 'b'}, - {"region",required_argument, 0, 'r'}, + {"base",no_argument, 0, 'b'}, + {"region",required_argument, 0, 'c'}, + {"reference",required_argument, 0, 'r'}, {"help", no_argument, 0, 'h'}, {"version", no_argument, 0, 'v'}, { NULL, 0, NULL, 0} @@ -232,7 +248,7 @@ void setup_options(int argc, char *argv[]){ int iarg = 0; //Iterate through options - while((iarg = getopt_long(argc, argv, "F:i:o:b:r:hv",long_opts, &index)) != -1){ + while((iarg = getopt_long(argc, argv, "F:i:o:c:r:bhv",long_opts, &index)) != -1){ switch(iarg){ case 'F': if(sscanf(optarg, "%i", &filter) != 1){ @@ -256,7 +272,7 @@ void setup_options(int argc, char *argv[]){ case 'v': print_version (0); break; - case 'r': + case 'c': region_store = optarg; //First check for a region format int res = check_region_string(region_store); @@ -265,30 +281,11 @@ void setup_options(int argc, char *argv[]){ print_usage(1); } break; + case 'r': + reference = optarg; + break; case 'b': - if(sscanf(optarg, "%c", &base) != 1){ - fprintf(stderr,"Error parsing -b|--base argument '%s'. Should be a single character > 0",optarg); - print_usage(1); - } - if(base != 'A' && base != 'C' && base != 'G' && base != 'T'){ - fprintf(stderr,"Single base %c provided must match [ACGT].\n",base); - print_usage(1); - }else{ - switch (base){ - case 'A': - base_bit = 1; - break; - case 'C': - base_bit = 2; - break; - case 'G': - base_bit = 4; - break; - case 'T': - base_bit = 8; - break; - } - } + is_base = 1; break; case '?': print_usage (1); @@ -304,7 +301,7 @@ void setup_options(int argc, char *argv[]){ print_usage(1); } - if(base_bit != 0 && region_store==NULL){ + if(is_base && region_store==NULL){ fprintf(stderr,"Option -r|--region must be used with the -b|--base option.\n"); print_usage(1); } @@ -313,20 +310,49 @@ void setup_options(int argc, char *argv[]){ } // callback for bam_plbuf_init() -static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data) -{ +static int perbase_pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data){ + tmpstructbase_t *tmp = (tmpstructbase_t*)data; + int pos = (int)position; + int coverage = n; + int base_coverage = 0; + int i; + for (i=0;ibase_bit) base_coverage++; + } + } + float result = 0; + if(base_coverage>0) result = (float)base_coverage / (float) coverage; + if(tmp->ltid != tid || tmp->lbaseprop != result || pos > tmp->lpos+1){ + //if(tmp->lpos > 0){ + uint32_t start = tmp->lstart; + uint32_t stop = (tmp->lpos +1); + float res = tmp->lbaseprop; + bwAddIntervals(tmp->out,&tmp->head->target_name[tmp->ltid],&start,&stop,&res,single); + //} + tmp->ltid = tid; + tmp->lstart = pos; + tmp->lbaseprop = result; + } + tmp->lpos = pos; + return 0; +} + +// callback for bam_plbuf_init() +static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data){ tmpstruct_t *tmp = (tmpstruct_t*)data; int pos = (int)position; int coverage = n; int i; for (i=0;i0 && bam_seqi(bam_get_seq(pl[i].b), pl[i].qpos)!=base_bit)) coverage--; + if (pl[i].is_del) coverage--; if (tmp->ltid != tid || tmp->lcoverage != coverage || pos > tmp->lpos+1) { if (tmp->lpos > 0 && tmp->lcoverage > 0){ uint32_t start = tmp->lstart; - uint32_t stop = tmp->lpos+1; - float cvg = tmp->lcoverage; + uint32_t stop = (tmp->lpos +1); + float cvg = (float)tmp->lcoverage; bwAddIntervals(tmp->out,&tmp->head->target_name[tmp->ltid],&start,&stop,&cvg,single); } tmp->ltid = tid; @@ -337,9 +363,28 @@ static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1 return 0; } +bigWigFile_t *initialise_bw_output(char *out_file, chromList_t *chromList){ + //Open output file + bigWigFile_t *fp = bwOpen(out_file, NULL, "w"); + check(fp, "Error opening %s for writing.",out_file); + //10 zoom levels + int res = bwCreateHdr(fp, 10); + check(!res,"Received error %d in bwCreateHeader for %s",res,out_file); + //Create the chromosome lists + fp->cl = bwCreateChromList(chromList->chrom, chromList->len, chromList->nKeys); + check(fp->cl != NULL, "Error generating chromlist as bw for %s.",out_file); + res = bwWriteHdr(fp); + check(!res,"Error %d writing bw header for %s.",res,out_file); + + return fp; +error: + return NULL; +} + int main(int argc, char *argv[]){ setup_options(argc, argv); tmpstruct_t tmp; + tmpstructbase_t *perbase = NULL; int no_of_regions = 0; int sq_lines = get_no_of_SQ_lines(input_file); FILE *fp_bed = NULL; @@ -413,7 +458,6 @@ int main(int argc, char *argv[]){ //Now create a bigwigfile //Open file as a bw file chromList_t *chromList = NULL; - bigWigFile_t *bw_out = NULL; bam_plp_t buf = NULL; bam1_t *b = NULL; hts_itr_t *iter = NULL; @@ -429,32 +473,58 @@ int main(int argc, char *argv[]){ int res = build_chromList_from_bam(chromList,input_file); check(res==1,"Error building chromList from bam header."); - //Open output file - bw_out = bwOpen(out_file, NULL, "w"); - check(bw_out!=NULL, "Error opening %s for writing.",out_file); //Initialise bw res = bwInit(1<<17); check(res==0,"Received an error in bwInit"); - //10 zoom levels - res = bwCreateHdr(bw_out, 10); - check(res==0,"Received error %d in bwCreateHeader",res); - //Create the chromosome lists - bw_out->cl = bwCreateChromList(chromList->chrom, chromList->len, chromList->nKeys); - check(bw_out->cl != NULL, "Error generating chromlist as bw."); - res = bwWriteHdr(bw_out); - check(res==0,"Error %d writing bw header.",res); + + if(!is_base){ + tmp.out = initialise_bw_output(out_file,chromList); + check(tmp.out!= NULL,"Error initialising bw output file %s.",out_file); + } tmp.beg = 0; tmp.end = 0x7fffffff; tmp.lstart = 0; tmp.lcoverage = 0; tmp.ltid = 0; tmp.lpos = 0; - tmp.out = bw_out; - //Open input file tmp.in = hts_open(input_file, "r"); check(tmp.in!=0,"Failed to open [CR|B]AM file '%s'.", input_file); + if(reference){ + int check = hts_set_fai_filename(tmp.in, reference); + check(check==0,"Error setting reference %s for hts file %s.",reference,input_file); + } tmp.head = sam_hdr_read(tmp.in); + check(tmp.head!=0,"Failed to read header from [CR|B]AM file '%s'.", input_file); + + if(is_base){ + uint8_t base_bit_list[] = {1,2,4,8}; + char *base_list[] = {"A","C","G","T"}; + perbase = malloc(4 * sizeof(tmpstructbase_t)); + check_mem(perbase); + int l=0; + for(l=0;l<4;l++){ + perbase[l].lstart = 0; + perbase[l].lcoverage = 0; + perbase[l].ltid = 0; + perbase[l].lpos = 0; + perbase[l].beg = 0; + perbase[l].end = 0x7fffffff; + perbase[l].lbaseprop = 0; + perbase[l].base_bit = base_bit_list[l]; + char *name = malloc(sizeof(char *) * (strlen(out_file)+3)); + check_mem(name); + memcpy(name,base_list[l],sizeof(base_list[l])); + strcat(name,"."); + strcat(name,out_file); + perbase[l].out = initialise_bw_output(name,chromList); + check(perbase[l].out != NULL,"Error initialising bw output file %s.",name); + free(name); + perbase[l].in = tmp.in; + perbase[l].head = tmp.head; + } + } + //Now we generate the bw info int i=0; for(i=0;i= 0) { if((b->core.flag & filter)>0) continue; //Skip if this is a filtered read @@ -474,7 +543,15 @@ int main(int argc, char *argv[]){ ret = bam_plp_push(buf, b); if (ret < 0) break; while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ + if(is_base){ + int b=0; + for(b=0;b<4;b++){ + perbase[b].idx = tmp.idx; + perbase_pileup_func(tid, pos, n_plp, plp, &perbase[b]); + } + }else{ pileup_func(tid, pos, n_plp, plp, &tmp); + } } } bam_plp_push(buf,0); @@ -483,19 +560,45 @@ int main(int argc, char *argv[]){ int n_plp, tid, pos; const bam_pileup1_t *plp; while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ - pileup_func(tid, pos, n_plp, plp, &tmp); + if(is_base){ + int b=0; + for(b=0;b<4;b++){ + perbase_pileup_func(tid, pos, n_plp, plp, &perbase[b]); + } + }else{ + pileup_func(tid, pos, n_plp, plp, &tmp); + } } uint32_t start = tmp.lstart; uint32_t stop = tmp.lpos+1; float cvg = tmp.lcoverage; - bwAddIntervals(tmp.out,&tmp.head->target_name[tmp.ltid],&start,&stop,&cvg,single); + if(is_base){ + int b=0; + for(b=0;b<4;b++){ + float result = perbase[b].lbaseprop; + uint32_t start = perbase[b].lstart; + uint32_t stop = perbase[b].lpos+1; + bwAddIntervals(perbase[b].out,&perbase[b].head->target_name[perbase[b].ltid],&start,&stop,&result,single); + } + }else{ + bwAddIntervals(tmp.out,&tmp.head->target_name[tmp.ltid],&start,&stop,&cvg,single); + } bam_plp_destroy(buf); bam_destroy1(b); sam_itr_destroy(iter); } - bwClose(bw_out); + + if(is_base){ + int b=0; + for(b=0;b<4;b++){ + bwClose(perbase[b].out); + } + }else{ + bwClose(tmp.out); + } + bwCleanup(); hts_close(tmp.in); int clean=0; @@ -504,9 +607,11 @@ int main(int argc, char *argv[]){ free(chromList->chrom[clean]); } } + free(chromList->chrom); free(chromList->len); free(chromList); hts_idx_destroy(tmp.idx); + if(is_base) free(perbase); bam_hdr_destroy(tmp.head); free(our_region_list); return 0; @@ -514,5 +619,6 @@ int main(int argc, char *argv[]){ error: if(our_region_list) free(our_region_list); if(fp_bed) fclose(fp_bed); + if(perbase) free(perbase); return -1; } From 991346e5909ab936befa4fb4f62e23048099518c Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Mon, 3 Oct 2016 16:36:32 +0100 Subject: [PATCH 3/8] Attempted conversion fo cgpBigWig --- .travis.yml | 6 + README.md | 17 +- bin/bamToBw.pl | 4 +- c/Makefile | 32 +-- c/bam2bedgraph.c | 228 ----------------- c/bam2bw.c | 624 --------------------------------------------- c/bwcat.c | 400 ----------------------------- lib/PCAP.pm | 12 + lib/PCAP/BigWig.pm | 16 +- setup.sh | 49 +--- 10 files changed, 48 insertions(+), 1340 deletions(-) delete mode 100644 c/bam2bedgraph.c delete mode 100644 c/bam2bw.c delete mode 100644 c/bwcat.c diff --git a/.travis.yml b/.travis.yml index c1e4294..eced1a8 100644 --- a/.travis.yml +++ b/.travis.yml @@ -29,6 +29,12 @@ language: perl perl: - "5.22" +before_script: + - git clone --depth 1 --single-branch --branch dev https://github.com/cancerit/cgpBigWig.git + - cd cgpBigWig + - ./setup.sh $HOME/wtsi-opt + - cd ../ + script: - ./setup.sh ~/wtsi-opt - ~/wtsi-opt/bin/samtools view # dump usage to show intact diff --git a/README.md b/README.md index f8e59dc..b270187 100644 --- a/README.md +++ b/README.md @@ -18,19 +18,22 @@ Please see the [wiki](https://github.com/ICGC-TCGA-PanCancer/PCAP-core/wiki) for --- ###Dependencies/Install -Some of the code included in this package has dependencies on external packages: - * [biobambam](https://github.com/gt1/biobambam) - * [bwa](https://github.com/lh3/bwa) - * [samtools](https://github.com/samtools/samtools) - * [kentUtils](https://github.com/ENCODE-DCC/kentUtils) +Please install the following before running `setup.sh`: + +* [cgpBigWig](https://github.com/cancerit/cgpBigWig/releases) + +Dependancies installed by `setup.sh`: + +* [biobambam](https://github.com/gt1/biobambam) +* [bwa](https://github.com/lh3/bwa) +* [samtools](https://github.com/samtools/samtools) And various perl modules. Please see the respective licence for each before use. -Please use `setup.sh` to install the dependencies. Please be aware that this expects basic C -compilation libraries and tools to be available, most are listed in `INSTALL`. +Please be aware that this expects basic C compilation libraries and tools to be available, most are listed in `INSTALL`. --- diff --git a/bin/bamToBw.pl b/bin/bamToBw.pl index bc86eb1..3b871b4 100755 --- a/bin/bamToBw.pl +++ b/bin/bamToBw.pl @@ -47,8 +47,10 @@ { my $options = setup(); + $options->{'ctg_lengths'} = PCAP::ref_lengths($options->{'reference'}.'.fai'); + my $threads = PCAP::Threaded->new($options->{'threads'}); - &PCAP::Threaded::disable_out_err if(exists $options->{'index'}); + &PCAP::Threaded::disable_out_err unless(exists $options->{'index'}); # register processes $threads->add_function('bamToBw', \&PCAP::BigWig::bamToBw); diff --git a/c/Makefile b/c/Makefile index 818c4b0..ce223f4 100644 --- a/c/Makefile +++ b/c/Makefile @@ -14,11 +14,6 @@ CFLAGS = -Wall HTSLOC?=$(HTSLIB) -#if test -z "$$LIBBIGWIG" -#LIBBIGWIGLOC?=$(LIBBIGWIG) -#echo "*******DEFINED" -#endif - HTSTMP?=./htslib_tmp prefix=?/usr/local/ @@ -26,11 +21,6 @@ prefix=?/usr/local/ OPTINC?= -I$(HTSLOC)/ INCLUDES= -I./ $(OPTINC) -rdynamic -ifdef LIBBIGWIG -CAT_INCLUDES= -I$(LIBBIGWIG) -CAT_LFLAGS= -L$(LIBBIGWIG) -endif - CAT_INCLUDES+= -I$(prefix)/include CAT_LFLAGS+= -L$(prefix)/lib @@ -64,10 +54,7 @@ MD := mkdir #Build target executable BAM_STATS_TARGET=../bin/bam_stats -CAT_TARGET=../bin/bwcat SQ_TARGET=../bin/reheadSQ -BAM2BG_TARGET=../bin/bam2bedgraph -BAM2BW_TARGET=../bin/bam2bw # # The following part of the makefile is generic; it can be used to @@ -80,20 +67,11 @@ BAM2BW_TARGET=../bin/bam2bw .NOTPARALLEL: test all: clean pre make_htslib_tmp $(BAM_STATS_TARGET) $(BAM2BG_TARGET) $(BAM2BW_TARGET) test remove_htslib_tmp $(CAT_TARGET) $(SQ_TARGET) - @echo bam_stats, reheadSQ, bwcat and bam2bw compiled. + @echo bam_stats and reheadSQ compiled. $(BAM_STATS_TARGET): $(OBJS) $(CC) $(CFLAGS) $(INCLUDES) -o $(BAM_STATS_TARGET) $(OBJS) $(LFLAGS) $(LIBS) ./bam_stats.c -$(CAT_TARGET): - $(CC) $(CAT_INCLUDES) $(CFLAGS) ./bwcat.c $(CAT_LFLAGS) -lBigWig -lz -lcurl -lm -lgnutls -ltasn1 -lhogweed -lnettle -lgmp -lp11-kit -o $(CAT_TARGET) - -$(BAM2BW_TARGET): - $(CC) $(CAT_INCLUDES) $(INCLUDES) $(CFLAGS) ./bam2bw.c $(CAT_LFLAGS) -lBigWig -lz -lcurl -lm -lgnutls -ltasn1 -lhogweed -lnettle -lgmp -lp11-kit $(LFLAGS) $(LIBS) -o $(BAM2BW_TARGET) - -$(BAM2BG_TARGET): $(OBJS) - $(CC) $(CFLAGS) $(INCLUDES) -o $(BAM2BG_TARGET) $(OBJS) $(LFLAGS) $(LIBS) ./bam2bedgraph.c - $(SQ_TARGET): $(CC) $(CFLAGS) ./reheadSQ.c -o $(SQ_TARGET) @@ -108,12 +86,6 @@ test: $(TESTS) coverage: CFLAGS += --coverage coverage: test -make_bam2bw: clean pre make_htslib_tmp $(BAM2BW_TARGET) remove_htslib_tmp - @echo $(BAM2BW_TARGET) done - -make_bigwigcat: $(CAT_TARGET) - @echo $(CAT_TARGET) done - make_htslib_tmp: $(MD) $(HTSTMP) #Do some magic to ensure we compile BAM_STATS with the static libhts.a rather than libhts.so @@ -140,7 +112,7 @@ valgrind: clean: @echo clean - $(RM) ./*.o *~ $(BAM_STATS_TARGET) $(SQ_TARGET) $(CAT_TARGET) $(BAM2BW_TARGET) $(BAM2BG_TARGET) ./tests/tests_log $(TESTS) ./*.gcda ./*.gcov ./*.gcno *.gcda *.gcov *.gcno ./tests/*.gcda ./tests/*.gcov ./tests/*.gcno + $(RM) ./*.o *~ $(BAM_STATS_TARGET) $(SQ_TARGET) ./tests/tests_log $(TESTS) ./*.gcda ./*.gcov ./*.gcno *.gcda *.gcov *.gcno ./tests/*.gcda ./tests/*.gcov ./tests/*.gcno -rm -rf $(HTSTMP) depend: $(SRCS) diff --git a/c/bam2bedgraph.c b/c/bam2bedgraph.c deleted file mode 100644 index 7b97790..0000000 --- a/c/bam2bedgraph.c +++ /dev/null @@ -1,228 +0,0 @@ -#include -#include -#include -#include -#include "htslib/sam.h" - -static char *input_file = NULL; -static char *output_file = NULL; -static char *region = NULL; -static int filter = 0; - -typedef struct { - uint32_t ltid; - int lstart,lcoverage,lpos,beg,end; - htsFile *in; - hts_idx_t *idx; - bam_hdr_t *head; - FILE *out; -} tmpstruct_t; - -void print_version (int exit_code){ - printf ("%s\n",VERSION); - exit(exit_code); -} - -int check_exist(char *fname){ - FILE *fp; - if((fp = fopen(fname,"r"))){ - fclose(fp); - return 1; - } - return 0; -} - -void print_usage (int exit_code){ - - printf ("Usage: bam2bedgraph -i input.[cr|b]am -o file [-r region] [-h] [-v]\n\n"); - printf ("Create a BEDGraph file of genomic coverage. BAM file must be sorted.\n"); - printf ("-i --input Path to bam/cram input file. [default: stdin]\n"); - printf ("-o --output File path for output. [default: stdout]\n\n"); - printf ("Optional:\n"); - printf ("-r --region Region in bam to access.\n"); - printf ("-f --filter Ignore reads with the filter flags [int].\n"); - printf ("Other:\n"); - printf ("-h --help Display this usage information.\n"); - printf ("-v --version Prints the version number.\n\n"); - exit(exit_code); -} - -void options(int argc, char *argv[]){ - const struct option long_opts[] = - { - {"version", no_argument, 0, 'v'}, - {"help",no_argument,0,'h'}, - {"input",required_argument,0,'i'}, - {"region",required_argument,0,'r'}, - {"filter",required_argument,0,'f'}, - {"output",required_argument,0,'o'}, - {"rna",no_argument,0, 'a'}, - { NULL, 0, NULL, 0} - - }; //End of declaring opts - - int index = 0; - int iarg = 0; - - //Iterate through options - while((iarg = getopt_long(argc, argv, "i:o:r:f:vh", long_opts, &index)) != -1){ - switch(iarg){ - case 'i': - input_file = optarg; - break; - - case 'o': - output_file = optarg; - break; - - case 'r': - region = optarg; - break; - - case 'f': - if(sscanf(optarg, "%i", &filter) != 1){ - printf("Error parsing -f argument '%s'. Should be an integer > 0",optarg); - print_usage(1); - } - break; - case 'h': - print_usage(0); - break; - - case 'v': - print_version(0); - break; - - case '?': - print_usage (1); - break; - - default: - print_usage (1); - - }; // End of args switch statement - - }//End of iteration through options - - //Do some checking to ensure required arguments were passed and are accessible files - if (input_file==NULL || strcmp(input_file,"/dev/stdin")==0) { - input_file = "-"; // htslib recognises this as a special case - } - if (strcmp(input_file,"-") != 0) { - if(check_exist(input_file) != 1){ - printf("Input file (-i) %s does not exist.\n",input_file); - print_usage(1); - } - } - if (output_file==NULL || strcmp(output_file,"/dev/stdout")==0) { - output_file = "-"; // we recognise this as a special case - } - - return; -} - -// callback for bam_plbuf_init() -static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data) -{ - tmpstruct_t *tmp = (tmpstruct_t*)data; - int pos = (int)position; - int coverage = n; - int i; - for (i=0;iltid != tid || tmp->lcoverage != coverage || pos > tmp->lpos+1) { - if (tmp->lpos > 0 && tmp->lcoverage > 0) - fprintf(tmp->out,"%s\t%d\t%d\t%d\n", tmp->head->target_name[tmp->ltid], tmp->lstart,tmp->lpos+1, tmp->lcoverage); - tmp->ltid = tid; - tmp->lstart = pos; - tmp->lcoverage = coverage; - } - tmp->lpos = pos; - return 0; -} - -int main(int argc, char *argv[]){ - options(argc, argv); - tmpstruct_t tmp; - bam_plp_t buf = NULL; - bam1_t *b = NULL; - hts_itr_t *iter = NULL; - tmp.beg = 0; tmp.end = 0x7fffffff; - tmp.lstart = 0; - tmp.lcoverage = 0; - tmp.ltid = 0; - tmp.lpos = 0; - - if (strcmp(output_file,"-")==0) { - tmp.out = stdout; - } else { - tmp.out = fopen(output_file,"w"); - } - if(tmp.out == NULL){ - fprintf(stderr,"Failed to open output file for %s writing.",output_file); - return 1; - } - tmp.in = hts_open(input_file, "r"); - if (tmp.in == 0) { - fprintf(stderr, "Fail to open [CR|B]AM file %s\n", input_file); - return 1; - } - tmp.head = sam_hdr_read(tmp.in); - buf = bam_plp_init(0,0); - //Iterate through each read in bam file. - b = bam_init1(); - if(region == NULL){ - tmp.idx=NULL; - int reto; - while((reto = sam_read1(tmp.in, tmp.head, b)) >= 0){ - if((b->core.flag & filter)>0) continue; //Skip if this is a filtered read - int ret, n_plp, tid, pos; - const bam_pileup1_t *plp; - ret = bam_plp_push(buf, b); - if (ret < 0) break; - while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0) - pileup_func(tid, pos, n_plp, plp, &tmp); - } - bam_plp_push(buf,0); - }else{ - - tmp.idx = sam_index_load(tmp.in,input_file); - if (tmp.idx == 0) { - fprintf(stderr, "Fail to open [CR|B]AM index for file %s\n", input_file); - return 1; - } - iter = sam_itr_querys(tmp.idx, tmp.head, region); - int result; - while ((result = sam_itr_next(tmp.in, iter, b)) >= 0) { - if((b->core.flag & filter)>0) continue; //Skip if this is a filtered read - int ret, n_plp, tid, pos; - const bam_pileup1_t *plp; - ret = bam_plp_push(buf, b); - if (ret < 0) break; - while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ - pileup_func(tid, pos, n_plp, plp, &tmp); - } - } - bam_plp_push(buf,0); - } - - //Check we've written everything... - int n_plp, tid, pos; - const bam_pileup1_t *plp; - while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ - pileup_func(tid, pos, n_plp, plp, &tmp); - } - - fprintf(tmp.out,"%s\t%d\t%d\t%d\n", tmp.head->target_name[tmp.ltid], tmp.lstart,tmp.lpos+1, tmp.lcoverage); - bam_plp_destroy(buf); - bam_destroy1(b); - fflush(tmp.out); - fclose(tmp.out); - if(iter) sam_itr_destroy(iter); - if(tmp.idx) hts_idx_destroy(tmp.idx); - if(tmp.in) hts_close(tmp.in); - if(tmp.head) bam_hdr_destroy(tmp.head); - return 0; -} - diff --git a/c/bam2bw.c b/c/bam2bw.c deleted file mode 100644 index 719bf67..0000000 --- a/c/bam2bw.c +++ /dev/null @@ -1,624 +0,0 @@ -#include -#include -#include -#include -#include -#include -#include "bigWig.h" -#include "htslib/hts.h" -#include "htslib/sam.h" -#include "dbg.h" - -char *out_file = "output.bam.bw"; -char *input_file = NULL; -char *region_store = NULL; -char **our_region_list = NULL; -char *reference = NULL; -uint32_t region_list_count = 0; -int is_regions_file = 0; -uint8_t is_base = 0; -int filter = 4; -char base = 0; -uint32_t single = 1; - -typedef struct { - uint32_t ltid; - int lstart,lcoverage,lpos,beg,end; - htsFile *in; - hts_idx_t *idx; - bam_hdr_t *head; - bigWigFile_t *out; - -} tmpstruct_t; - -typedef struct { - uint32_t ltid; - int lstart,lcoverage,lpos,beg,end; - float lbaseprop; - htsFile *in; - hts_idx_t *idx; - bam_hdr_t *head; - bigWigFile_t *out; - uint8_t base_bit; -} tmpstructbase_t; - -void print_version (int exit_code){ - printf ("%s\n",VERSION); - exit(exit_code); -} - -int line_count (char *file_path){ - FILE *f = fopen(file_path,"r"); - int line_count = 0; - check(f != NULL, "Error opening file '%s' to count lines.",file_path); - char rd[512]; - while(fgets(rd, 512, f) != NULL){ - line_count++; - } - fclose(f); - return line_count; -error: - if(f) fclose(f); - return -1; -} - -int check_exist(char *fname){ - FILE *fp; - if((fp = fopen(fname,"r"))){ - fclose(fp); - return 1; - } - return 0; -} - -void print_usage (int exit_code){ - printf("Usage: bam2bw -i input.[b|cr]am -o output.bw\n"); - printf("bam2bw can be used to generate a bw file of coverage from a [cr|b]am file or a per base proportion bigwig file over a specified region.\n\n"); - printf("-i --input [file] Path to the input [b|cr]am file.\n"); - printf("-F --filter [int] SAM flags to filter. [default: %d]\n",filter); - printf("-o --outfile [file] Path to the output .bw file produced. Per base results wiillbe output as four bw files [ACGT].outputname.bw [default:'%s']\n\n",out_file); - printf("Optional: \n"); - printf("-b --perbase To produce per base coverage bigwig files [ACTG]. Must be used in conjunction with -r|--region\n"); - printf("-c --region [file] A samtools style region (contig:start-stop) or a bed file of regions over which to produce the bigwig file\n"); - printf("-r --reference [file] Path to reference genome.fa file (required for cram if ref_path cannot be resolved)\n\n"); - printf ("Other:\n"); - printf("-h --help Display this usage information.\n"); - printf("-v --version Prints the version number.\n\n"); - - exit(exit_code); -} - -int check_region_string(char *region){ - if(check_exist(region)==1){ - is_regions_file = 1; - return 1; - }else{ - //Parse this as a region - int beg = 0; - int end = 0; - const char *res = hts_parse_reg(region, &beg, &end); - if(res) return 1; - } - return -1; -} - -int get_int_length(int input){ - return (input == 0 ? 1 : (int)(log10(input)+1)); -} - -int get_no_of_SQ_lines(char *bam_loc){ - htsFile *bam = NULL; - char *line = NULL; - bam_hdr_t *header = NULL; - bam = hts_open(bam_loc, "r"); - check(bam != 0,"Bam file %s failed to open to read header.",bam_loc); - header = sam_hdr_read(bam); - char *head_txt = header->text; - line = strtok(head_txt,"\n"); - int count = 0; - while(line != NULL){ - //Check for a read group line - if(strncmp(line,"@SQ",3)==0){ - count++; - }//End of if this is an SQ line - line = strtok(NULL,"\n"); - } - free(line); - hts_close(bam); - bam_hdr_destroy(header); - return count; -error: - if(line) free(line); - if(bam) hts_close(bam); - if(header) bam_hdr_destroy(header); - return -1; -} - -int parse_SQ_line(char *line, char *name, int *length){ - char *tag = NULL; - char *tmp = NULL; - char *ptr = NULL; - int nom = 0; - int len = 0; - tag = strtok_r(line,"\t",&ptr); - while(tag != NULL){ - int chk=0; - tmp = malloc(sizeof(char) * 512); - check_mem(tmp); - chk = sscanf(tag,"SN:%[^\t\n]",tmp); - if(chk>0){ - strcpy(name,tmp); - tag = strtok_r(NULL,"\t",&ptr); - nom = 1; - continue; - } - chk = sscanf(tag,"LN:%d",length); - if(chk>0){ - len = 1; - tag = strtok_r(NULL,"\t",&ptr); - continue; - } - tag = strtok_r(NULL,"\t",&ptr); - } - if(tmp) free(tmp); - if(tag) free(tag); - - if(nom && len) { - return 1; - }else{ - return -1; - } -error: - if(tmp) free(tmp); - if(tag) free(tag); - return -1; -} - -int build_chromList_from_bam(chromList_t *chromList ,char *bam_loc){ - htsFile *bam = NULL; - char *line = NULL; - char *tmp = NULL; - bam_hdr_t *header = NULL; - char *ptr; - bam = hts_open(bam_loc, "r"); - check(bam != 0,"Bam file %s failed to open to read header.",bam_loc); - header = sam_hdr_read(bam); - char *head_txt = header->text; - line = strtok(head_txt,"\n"); - int i = 0; - while(line != NULL){ - //Check for a read group line - if(strncmp(line,"@SQ",3)==0){ - char *tag = strtok_r(line,"\t",&ptr); - while(tag != NULL){ - int chk=0; - tmp = malloc(sizeof(char) * 512); - check_mem(tmp); - chk = sscanf(tag,"SN:%[^\t\n]",tmp); - if(chk>0){ - chromList->chrom[i] = malloc(sizeof(char) * (strlen(tmp)+1)); - check_mem(chromList->chrom[i]); - strcpy(chromList->chrom[i],tmp); - tag = strtok_r(NULL,"\t",&ptr); - free(tmp); - continue; - } - free(tmp); - uint32_t tmpint = 0; - chk = sscanf(tag,"LN:%" SCNu32 "",&tmpint); - if(chk>0){ - chromList->len[i] = tmpint; - tag = strtok_r(NULL,"\t",&ptr); - continue; - } - tag = strtok_r(NULL,"\t",&ptr); - }//End of SQ line tags - i++; - }//End of if this is an SQ line - line = strtok(NULL,"\n"); - } - free(line); - bam_hdr_destroy(header); - hts_close(bam); - return 1; -error: - if(line) free(line); - if(bam) hts_close(bam); - if(header) bam_hdr_destroy(header); - if(tmp) free (tmp); - return -1; -} - -void setup_options(int argc, char *argv[]){ - const struct option long_opts[] = - { - {"input", required_argument, 0, 'i'}, - {"filter", required_argument, 0, 'F'}, - {"outfile",required_argument, 0, 'o'}, - {"base",no_argument, 0, 'b'}, - {"region",required_argument, 0, 'c'}, - {"reference",required_argument, 0, 'r'}, - {"help", no_argument, 0, 'h'}, - {"version", no_argument, 0, 'v'}, - { NULL, 0, NULL, 0} - - }; //End of declaring opts - - int index = 0; - int iarg = 0; - - //Iterate through options - while((iarg = getopt_long(argc, argv, "F:i:o:c:r:bhv",long_opts, &index)) != -1){ - switch(iarg){ - case 'F': - if(sscanf(optarg, "%i", &filter) != 1){ - fprintf(stderr,"Error parsing -F|--filter argument '%s'. Should be an integer > 0",optarg); - print_usage(1); - } - break; - case 'i': - input_file = optarg; - if(check_exist(input_file) != 1){ - fprintf(stderr,"Input bam file %s does not appear to exist.\n",input_file); - print_usage(1); - } - break; - case 'o': - out_file = optarg; - break; - case 'h': - print_usage (0); - break; - case 'v': - print_version (0); - break; - case 'c': - region_store = optarg; - //First check for a region format - int res = check_region_string(region_store); - if(res<0){ - fprintf(stderr,"Region %s is not in correct format or not an existing bed file.\n",region_store); - print_usage(1); - } - break; - case 'r': - reference = optarg; - break; - case 'b': - is_base = 1; - break; - case '?': - print_usage (1); - break; - default: - print_usage (0); - }; // End of args switch statement - - }//End of iteration through options - - if(input_file==NULL){ - fprintf(stderr,"Required option -i|--input-bam not defined.\n"); - print_usage(1); - } - - if(is_base && region_store==NULL){ - fprintf(stderr,"Option -r|--region must be used with the -b|--base option.\n"); - print_usage(1); - } - - return; -} - -// callback for bam_plbuf_init() -static int perbase_pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data){ - tmpstructbase_t *tmp = (tmpstructbase_t*)data; - int pos = (int)position; - int coverage = n; - int base_coverage = 0; - int i; - for (i=0;ibase_bit) base_coverage++; - } - } - float result = 0; - if(base_coverage>0) result = (float)base_coverage / (float) coverage; - if(tmp->ltid != tid || tmp->lbaseprop != result || pos > tmp->lpos+1){ - //if(tmp->lpos > 0){ - uint32_t start = tmp->lstart; - uint32_t stop = (tmp->lpos +1); - float res = tmp->lbaseprop; - bwAddIntervals(tmp->out,&tmp->head->target_name[tmp->ltid],&start,&stop,&res,single); - //} - tmp->ltid = tid; - tmp->lstart = pos; - tmp->lbaseprop = result; - } - tmp->lpos = pos; - return 0; -} - -// callback for bam_plbuf_init() -static int pileup_func(uint32_t tid, uint32_t position, int n, const bam_pileup1_t *pl, void *data){ - tmpstruct_t *tmp = (tmpstruct_t*)data; - int pos = (int)position; - int coverage = n; - int i; - for (i=0;iltid != tid || tmp->lcoverage != coverage || pos > tmp->lpos+1) { - if (tmp->lpos > 0 && tmp->lcoverage > 0){ - uint32_t start = tmp->lstart; - uint32_t stop = (tmp->lpos +1); - float cvg = (float)tmp->lcoverage; - bwAddIntervals(tmp->out,&tmp->head->target_name[tmp->ltid],&start,&stop,&cvg,single); - } - tmp->ltid = tid; - tmp->lstart = pos; - tmp->lcoverage = coverage; - } - tmp->lpos = pos; - return 0; -} - -bigWigFile_t *initialise_bw_output(char *out_file, chromList_t *chromList){ - //Open output file - bigWigFile_t *fp = bwOpen(out_file, NULL, "w"); - check(fp, "Error opening %s for writing.",out_file); - //10 zoom levels - int res = bwCreateHdr(fp, 10); - check(!res,"Received error %d in bwCreateHeader for %s",res,out_file); - //Create the chromosome lists - fp->cl = bwCreateChromList(chromList->chrom, chromList->len, chromList->nKeys); - check(fp->cl != NULL, "Error generating chromlist as bw for %s.",out_file); - res = bwWriteHdr(fp); - check(!res,"Error %d writing bw header for %s.",res,out_file); - - return fp; -error: - return NULL; -} - -int main(int argc, char *argv[]){ - setup_options(argc, argv); - tmpstruct_t tmp; - tmpstructbase_t *perbase = NULL; - int no_of_regions = 0; - int sq_lines = get_no_of_SQ_lines(input_file); - FILE *fp_bed = NULL; - if(region_store){ - if(is_regions_file==1){ - //If we have a bedfile - no_of_regions = line_count(region_store); - check(no_of_regions>0,"Error counting entries in region file %s\n",region_store); - our_region_list = calloc(no_of_regions, sizeof(char *)); - check_mem(region_list); - int i=0; - fp_bed = fopen(region_store,"r"); - char line[512]; - while(fgets(line,512,fp_bed)){ - char *contig = malloc(sizeof(char)*256); - check_mem(contig); - int beg,end; - int chk = sscanf(line,"%s\t%d\t%d",contig,&beg,&end); - check(chk==3,"Error reading line '%s' from regions bed file.",line); - char *region = malloc(sizeof(char) * (strlen(contig)+get_int_length(beg)+get_int_length(end))+3); - check_mem(region); - chk = sprintf(region,"%s:%d-%d",contig,beg+1,end); //+1 to beginning as this is bed - check(chk==((strlen(contig)+get_int_length(beg)+get_int_length(end))+2),"Error creating region line from bed entry '%s'.",line); - our_region_list[i] = region; - i++; - } - fclose(fp_bed); - }else{ - //If we have a single region.... - no_of_regions = 1; - our_region_list = calloc(no_of_regions, sizeof(char *)); - check_mem(region_list); - our_region_list[0] = region_store; - } - }else{ - //If not... - //Build a list of regions from the header of the bamfile - no_of_regions = sq_lines; - our_region_list = calloc(no_of_regions, sizeof(char *)); - htsFile *bam = NULL; - char *line = NULL; - bam = hts_open(input_file, "r"); - check(bam != 0,"Bam file %s failed to open to read header.",input_file); - bam_hdr_t *header = sam_hdr_read(bam); - char *head_txt = header->text; - line = strtok(head_txt,"\n"); - int i=0; - while(line != NULL){ - //Check for a read group line - if(strncmp(line,"@SQ",3)==0){ - char *contig = malloc(sizeof(char) * 256); - int end,chk; - chk = parse_SQ_line(line,contig,&end); - check(chk==1,"Error parsing SQ line %s.\n",line); - char *region = malloc(sizeof(char) * (strlen(contig)+get_int_length(end))+4); - check_mem(region); - chk = sprintf(region,"%s:%d-%d",contig,1,end); - check(chk==((strlen(contig)+get_int_length(end))+3),"Error creating region line from bed entry '%s'.",line); - our_region_list[i] = region; - i++; - }//End of if this is an SQ line - line = strtok(NULL,"\n"); - } - bam_hdr_destroy(header); - free(line); - hts_close(bam); - } - - check(no_of_regions>0,"Error evaluating regions to parse. None found."); - - //Now create a bigwigfile - //Open file as a bw file - chromList_t *chromList = NULL; - bam_plp_t buf = NULL; - bam1_t *b = NULL; - hts_itr_t *iter = NULL; - //Generate the list of chromosomes using the bam header. - chromList = calloc(1, sizeof(chromList_t)); - check_mem(chromList); - chromList->nKeys = sq_lines; - chromList->chrom = calloc(sq_lines,sizeof(char *)); - check_mem(chromList->chrom); - chromList->len = calloc(sq_lines,sizeof(uint32_t)); - check_mem(chromList->len); - //Iterate through the header of the bam file and get all contigs. - int res = build_chromList_from_bam(chromList,input_file); - check(res==1,"Error building chromList from bam header."); - - //Initialise bw - res = bwInit(1<<17); - check(res==0,"Received an error in bwInit"); - - if(!is_base){ - tmp.out = initialise_bw_output(out_file,chromList); - check(tmp.out!= NULL,"Error initialising bw output file %s.",out_file); - } - - tmp.beg = 0; tmp.end = 0x7fffffff; - tmp.lstart = 0; - tmp.lcoverage = 0; - tmp.ltid = 0; - tmp.lpos = 0; - //Open input file - tmp.in = hts_open(input_file, "r"); - check(tmp.in!=0,"Failed to open [CR|B]AM file '%s'.", input_file); - if(reference){ - int check = hts_set_fai_filename(tmp.in, reference); - check(check==0,"Error setting reference %s for hts file %s.",reference,input_file); - } - tmp.head = sam_hdr_read(tmp.in); - check(tmp.head!=0,"Failed to read header from [CR|B]AM file '%s'.", input_file); - - if(is_base){ - uint8_t base_bit_list[] = {1,2,4,8}; - char *base_list[] = {"A","C","G","T"}; - perbase = malloc(4 * sizeof(tmpstructbase_t)); - check_mem(perbase); - int l=0; - for(l=0;l<4;l++){ - perbase[l].lstart = 0; - perbase[l].lcoverage = 0; - perbase[l].ltid = 0; - perbase[l].lpos = 0; - perbase[l].beg = 0; - perbase[l].end = 0x7fffffff; - perbase[l].lbaseprop = 0; - perbase[l].base_bit = base_bit_list[l]; - char *name = malloc(sizeof(char *) * (strlen(out_file)+3)); - check_mem(name); - memcpy(name,base_list[l],sizeof(base_list[l])); - strcat(name,"."); - strcat(name,out_file); - perbase[l].out = initialise_bw_output(name,chromList); - check(perbase[l].out != NULL,"Error initialising bw output file %s.",name); - free(name); - perbase[l].in = tmp.in; - perbase[l].head = tmp.head; - } - } - - //Now we generate the bw info - int i=0; - for(i=0;i= 0) { - if((b->core.flag & filter)>0) continue; //Skip if this is a filtered read - int ret, n_plp, tid, pos; - const bam_pileup1_t *plp; - ret = bam_plp_push(buf, b); - if (ret < 0) break; - while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ - if(is_base){ - int b=0; - for(b=0;b<4;b++){ - perbase[b].idx = tmp.idx; - perbase_pileup_func(tid, pos, n_plp, plp, &perbase[b]); - } - }else{ - pileup_func(tid, pos, n_plp, plp, &tmp); - } - } - } - bam_plp_push(buf,0); - - //Check we've written everything... - int n_plp, tid, pos; - const bam_pileup1_t *plp; - while ((plp = bam_plp_next(buf, &tid, &pos, &n_plp)) != 0){ - if(is_base){ - int b=0; - for(b=0;b<4;b++){ - perbase_pileup_func(tid, pos, n_plp, plp, &perbase[b]); - } - }else{ - pileup_func(tid, pos, n_plp, plp, &tmp); - } - } - uint32_t start = tmp.lstart; - uint32_t stop = tmp.lpos+1; - float cvg = tmp.lcoverage; - if(is_base){ - int b=0; - for(b=0;b<4;b++){ - float result = perbase[b].lbaseprop; - uint32_t start = perbase[b].lstart; - uint32_t stop = perbase[b].lpos+1; - bwAddIntervals(perbase[b].out,&perbase[b].head->target_name[perbase[b].ltid],&start,&stop,&result,single); - } - }else{ - bwAddIntervals(tmp.out,&tmp.head->target_name[tmp.ltid],&start,&stop,&cvg,single); - } - - bam_plp_destroy(buf); - bam_destroy1(b); - sam_itr_destroy(iter); - } - - - if(is_base){ - int b=0; - for(b=0;b<4;b++){ - bwClose(perbase[b].out); - } - }else{ - bwClose(tmp.out); - } - - bwCleanup(); - hts_close(tmp.in); - int clean=0; - for(clean=0; cleanchrom){ - free(chromList->chrom[clean]); - } - } - free(chromList->chrom); - free(chromList->len); - free(chromList); - hts_idx_destroy(tmp.idx); - if(is_base) free(perbase); - bam_hdr_destroy(tmp.head); - free(our_region_list); - return 0; - -error: - if(our_region_list) free(our_region_list); - if(fp_bed) fclose(fp_bed); - if(perbase) free(perbase); - return -1; -} diff --git a/c/bwcat.c b/c/bwcat.c deleted file mode 100644 index 361ca1b..0000000 --- a/c/bwcat.c +++ /dev/null @@ -1,400 +0,0 @@ -#include -#include -#include -#include -#include -#include -#include -#include "bigWig.h" - -char *out_file = "concatenated.bw"; -char *fai = NULL; -char *path = NULL; -char *contig_input = NULL; -int ign_count = 0; - -int check_exist(char *fname){ - FILE *fp; - if((fp = fopen(fname,"r"))){ - fclose(fp); - return 1; - } - return 0; -} - -void print_version (int exit_code){ - printf ("%s\n",VERSION); - exit(exit_code); -} - -void print_usage (int exit_code){ - printf("Usage: bigWigCat -f genome.fai -o output.bw\n\n"); - printf("-p --input-path [dir] Path to the input bigwig files named [path]/.bw\n"); - printf("-f --fasta-index [file] Fasta index file (.fai)\n"); - printf("-o --outfile [file] Path to the output .bw file produced. [default:'%s']\n",out_file); - printf("-g --ignore-contigs [comma separated list] Comma separated list of contigs to ignore.\n\n"); - printf ("Other:\n"); - printf ("-h --help Display this usage information.\n"); - printf ("-v --version Prints the version number.\n\n"); - - exit(exit_code); -} - -int line_count(char *file){ - FILE *in = fopen(file,"r"); - if(in==NULL){ - fprintf(stderr,"Error opening input file %s for line count:%d.\n",file,errno); - return -1; - } - char line [ 5000 ]; - int line_count=0; - while ( fgets(line,sizeof(line),in) != NULL ){ - line_count++; - } - //Close input file - fclose(in); - return line_count; -} - -void setup_options(int argc, char *argv[]){ - const struct option long_opts[] = - { - {"input-path", required_argument, 0, 'p'}, - {"fasta-index", required_argument, 0, 'f'}, - {"outfile",required_argument,0,'o'}, - {"ignore-contigs",required_argument,0,'g'}, - {"help", no_argument, 0, 'h'}, - {"version", no_argument, 0, 'v'}, - { NULL, 0, NULL, 0} - }; //End of declaring opts - - int index = 0; - int iarg = 0; - - //Iterate through options - while((iarg = getopt_long(argc, argv, "f:o:g:p:hv",long_opts, &index)) != -1){ - switch(iarg){ - case 'p': - path = optarg; - break; - case 'f': - fai = optarg; - break; - case 'o': - out_file = optarg; - break; - case 'h': - print_usage (0); - break; - case 'v': - print_version (0); - break; - case 'g': - contig_input = optarg; - break; - case '?': - print_usage (1); - break; - default: - print_usage (0); - }; // End of args switch statement - - }//End of iteration through options - - //Do some checking to ensure required arguments were passed and are accessible files - if(check_exist(fai) != 1){ - fprintf(stderr,"Fasta index (.fai) file %s does not appear to exist.\n",fai); - print_usage(1); - } -} - -char *build_regex_string(char *input){ - char *new = NULL; - new = malloc(sizeof(char) * (strlen(input) + 50)); - int m=0; - new[0] = '^'; - int pos = 1; - for(m=0;mnKeys = contig_count; - chromList->chrom = malloc(contig_count * sizeof(char *)); - if(!chromList->chrom){ - fprintf(stderr,"Error assigning memory for chromList chrom\n"); - goto error; - } - chromList->len = malloc(contig_count * sizeof(uint32_t)); - if(!chromList->len){ - fprintf(stderr,"Error assigning memory for chromList len\n"); - goto error; - } - fileList=malloc(contig_count * sizeof(char *)); - if(!fileList){ - fprintf(stderr,"Error assigning memory for fileList\n"); - goto error; - } - - idx = fopen(fai,"r"); - if(idx==NULL){ - fprintf(stderr,"Error opening input fai file %s for reading:%d.\n",fai,errno); - } - - //Build list of chr names and filenames from fasta index - char line [ 5000 ]; - int i=0; - while ( fgets(line,sizeof(line),idx) != NULL ){ - char contig [ 4000 ]; - char filePath [5000] = ""; - uint32_t len; - int chk = sscanf(line, "%s\t%d", contig,&len); - if(chk<=0){ - fprintf(stderr,"Error fetching contig from fai at line %s\n",line); - } - chromList->chrom[i] = malloc(sizeof(char) * (strlen(contig)+1)); - if(!chromList->chrom[i]){ - fprintf(stderr,"Error allocating memory for chromList->chrom[%d]\n",i); - goto error; - } - strcpy(chromList->chrom[i],contig); - chromList->len[i] = len; - //fileList[i] = - strcpy(filePath,path); - strcat(filePath,"/"); - strcat(filePath,contig); - strcat(filePath,".bw"); - fileList[i] = malloc(sizeof(char) * (strlen(filePath)+1)); - if(!fileList[i]){ - fprintf(stderr,"Error allocating memory for fileList[%d]\n",i); - goto error; - } - strcpy(fileList[i],filePath); - i++; - } - //Close input file - fclose(idx); - - ignore = parse_contig_list(contig_input,chromList->chrom,chromList->nKeys); - - //Open output file - bw_out = bwOpen(out_file, NULL, "w"); - if(!bw_out) { - fprintf(stderr, "An error occurred while opening %s for writing\n",out_file); - return 1; - } - - int j=0; - int first=1; - for(j=0;jnKeys;j++){//Iterate through each contig and access file. - - if(ignore != NULL && check_ignore(ignore,ign_count,chromList->chrom[j])){ - continue; - } - //Open file as a bw file - bw_in = NULL; - //Initialize enough space to hold 128KiB (1<<17) of data at a time - if(bwInit(1<<17) != 0) { - fprintf(stderr, "Received an error in bwInit\n"); - return 1; - } - if(check_exist(fileList[j])!=1){ - fprintf(stderr,"BW file for reading '%s' doesn't seem to exist\n",fileList[j]); - } - - bw_in = bwOpen(fileList[j], NULL, "r"); - if(!bw_in) { - fprintf(stderr, "Error opening bw file %s\n",fileList[j]); - goto error; - } - - //Write the header to file - if(first==1){ - if(bwCreateHdr(bw_out, bw_in->hdr->nLevels)){ - fprintf(stderr,"Error creating header for output bw file"); - goto error; - } - bw_out->cl = bwCreateChromList(chromList->chrom, chromList->len, chromList->nKeys); - if(!bw_out->cl){ - fprintf(stderr,"Error creating chromlist for output fo file"); - goto error; - } - if(bwWriteHdr(bw_out)){ - fprintf(stderr,"Error writing header to file\n"); - goto error; - } - first=0; - } - - intervals = NULL; - intervals = bwGetOverlappingIntervals(bw_in, chromList->chrom[j], 0, chromList->len[j]); - if(!intervals){ - fprintf(stderr,"Error retrieving intervals for %s:%d-%d\n",chromList->chrom[j], 0, chromList->len[j]); - goto error; - } - - if(intervals->l){ - chrList = NULL; - chrList = malloc(intervals->l * sizeof(char *)); - if(!chrList){ - fprintf(stderr,"Error allocating memory for chromList\n"); - goto error; - } - uint64_t k=0; - for(k=0;kl;k++){ - chrList[k] = chromList->chrom[j]; - } - if(bwAddIntervals(bw_out,chrList,intervals->start,intervals->end,intervals->value,intervals->l)){ - fprintf(stderr,"Error adding %d intervals for chromosome %s.\n",intervals->l,chromList->chrom[j]); - goto error; - } - free(chrList); - } - bwDestroyOverlappingIntervals(intervals); - bwClose(bw_in); - - }//End of iteration through each contig - - bwClose(bw_out); - bwCleanup(); - int clean=0; - for(clean=0; cleannKeys; clean++){ - if(chromList->chrom){ - free(chromList->chrom[clean]); - } - if(fileList){ - free(fileList[clean]); - } - } - free(chromList->len); - free(chromList); - return 0; - -error: - fclose(idx); - bwClose(bw_out); - if(intervals) bwDestroyOverlappingIntervals(intervals); - if(bw_in){ - bwClose(bw_in); - bwCleanup(); - } - //TODO cleanup chromList object - if(chromList){ - int clean=0; - for(clean=0; cleannKeys; clean++){ - if(chromList->chrom){ - free(chromList->chrom[clean]); - } - if(fileList){ - free(fileList[clean]); - } - } - free(chromList->len); - free(chromList); - } - return 1; - -} \ No newline at end of file diff --git a/lib/PCAP.pm b/lib/PCAP.pm index 4c51aa7..730103b 100644 --- a/lib/PCAP.pm +++ b/lib/PCAP.pm @@ -25,6 +25,7 @@ use Const::Fast qw(const); use base 'Exporter'; use FindBin qw($Bin); use File::Which qw(which); +use autodie qw(:all); our $VERSION = '3.1.0'; our @EXPORT = qw($VERSION _which); @@ -106,6 +107,17 @@ sub _which { return $path; } +sub ref_lengths { + my $fai_file = shift; + my %ctg_lengths; + open my $FAI, '<', $fai_file; + while(my $l = <$FAI>) { + my ($ctg, $len) = split /\t/, $l; + $ctg_lengths{$ctg} = $len; + } + return \%ctg_lengths; +} + 1; __END__ diff --git a/lib/PCAP/BigWig.pm b/lib/PCAP/BigWig.pm index b2ac2d7..93fd52a 100644 --- a/lib/PCAP/BigWig.pm +++ b/lib/PCAP/BigWig.pm @@ -48,15 +48,13 @@ sub bamToBw { my $outfile = q{'}.File::Spec->catfile($options->{'tmp'}, $seq.'.bw').q{'}; - my $command = q{bash -c "set pipefail; }; - $command .= _which('bam2bedgraph'); - $command .= q{ -f }.$filter; - $command .= q{ -r }.$seq; + my $command = _which('bam2bw'); + $command .= q{ -F }.$filter; + $command .= q{ -z}; + $command .= sprintf q{ -c %s:1-%d}, $seq, $options->{'ctg_lengths'}->{$seq}; $command .= q{ -i }.$options->{'bam'}; - $command .= ' | '; - $command .= _which('wigToBigWig'); - $command .= ' -fixedSummaries -keepAllChromosomes stdin '.$options->{'reference'}.'.fai '.$outfile; - $command .= q{"}; + $command .= q{ -o }.$outfile; + $command .= q{ -r }.$options->{'reference'}; PCAP::Threaded::external_process_handler(File::Spec->catdir($tmp, 'logs'), $command, $index); @@ -72,7 +70,7 @@ sub generateBw { my $outfile = File::Spec->catfile($options->{'outdir'}, $options->{'sample'}.'.bw'); - my $command = sprintf '%s -p %s -f %s -o %s', _which('bwcat'), + my $command = sprintf '%s -p %s -f %s -o %s', _which('bwjoin'), $options->{'tmp'}, $options->{'reference'}.'.fai', $outfile; diff --git a/setup.sh b/setup.sh index 1ad5efd..d3aafa2 100755 --- a/setup.sh +++ b/setup.sh @@ -10,13 +10,9 @@ SOURCE_HTSLIB="https://github.com/samtools/htslib/releases/download/1.3.1/htslib # Bio::DB::HTS SOURCE_BIOBDHTS="https://github.com/Ensembl/Bio-HTS/archive/2.3.tar.gz" -# for bigwig -SOURCE_JKENT_BIN="https://github.com/ENCODE-DCC/kentUtils/raw/master/bin/linux.x86_64" # for Bio::DB::BigWig SOURCE_KENTSRC="ftp://ftp.sanger.ac.uk/pub/cancer/legacy-dependancies/jksrc.v334.zip" SOURCE_BIGFILE="http://www.cpan.org/authors/id/L/LD/LDS/Bio-BigFile-1.07.tar.gz" -# for fast merging of per-chr BW files -SOURCE_LIB_BW="https://github.com/dpryan79/libBigWig/archive/b36da5a06bffcc1b33c369e078b82f84625fd212.tar.gz" # for biobambam SOURCE_BBB_BIN_DIST="https://github.com/gt1/biobambam2/releases/download/2.0.54-release-20160802163650/biobambam2-2.0.54-release-20160802163650-x86_64-etch-linux-gnu.tar.gz" @@ -92,6 +88,14 @@ export PATH="$INST_PATH/bin:$PATH" SETUP_DIR=$INIT_DIR/install_tmp mkdir -p $SETUP_DIR +# check for cgpBigWig +if [ -e "$INST_PATH/bin/bwjoin" ]; then + echo -e "\tcgpBigWig installation found"; +else + echo -e "\tERROR: Please see README.md and install cgpBigWig"; + exit 1 +fi + ## grab cpanm and stick in workspace, then do a self upgrade into bin: get_file $SETUP_DIR/cpanm https://cpanmin.us/ perl $SETUP_DIR/cpanm -l $INST_PATH App::cpanminus @@ -155,25 +159,6 @@ else touch $SETUP_DIR/biohts.success fi - -echo -n "Building jkentUtils ..." -if [ -e $SETUP_DIR/jkentUtils.success ]; then - echo " previously installed ..."; -else - echo - cd $SETUP_DIR - if [[ `uname -m` == x86_64 ]] ; then - get_file $INST_PATH/bin/wigToBigWig $SOURCE_JKENT_BIN/wigToBigWig - chmod +x $INST_PATH/bin/wigToBigWig - touch $SETUP_DIR/jkentUtils.success - else - if [ ! -e $INST_DIR/bin/wigToBigWig ]; then - echo "Binaries only available for x86_64, please install wigToBigWig from kentUtils: https://github.com/ENCODE-DCC/kentUtils" - exit 1 - fi - fi -fi - echo -n "Building htslib ..." if [ -e $SETUP_DIR/htslib.success ]; then echo " previously installed ..."; @@ -216,21 +201,6 @@ else echo "samtools - No change between PCAP versions" fi -echo -n "Building libBigWig ..." -if [ -e $SETUP_DIR/libBigWig.success ]; then - echo " previously installed ..."; -else - echo - cd $SETUP_DIR - get_distro "libBigWig" $SOURCE_LIB_BW - mkdir -p libBigWig - tar --strip-components 1 -C libBigWig -zxf libBigWig.tar.gz - make -C libBigWig -j$CPU install prefix=$INST_PATH - rm -f $INST_PATH/lib/libBigWig.so - rm -f libBigWig.tar.gz - touch $SETUP_DIR/libBigWig.success -fi - cd $SETUP_DIR if [[ ",$COMPILE," == *,bwa,* ]] ; then echo -n "Building BWA ..." @@ -316,10 +286,7 @@ else make -C c clean env HTSLIB=$SETUP_DIR/htslib make -C c -j$CPU prefix=$INST_PATH cp bin/bam_stats $INST_PATH/bin/. - cp bin/bwcat $INST_PATH/bin/. cp bin/reheadSQ $INST_PATH/bin/. - cp bin/bam2bedgraph $INST_PATH/bin/. - cp bin/bam2bw $INST_PATH/bin/. touch $SETUP_DIR/bam_stats.success make -C c clean fi From cfd4f47aa2f4458b07cf49a38519208754d339cc Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Mon, 3 Oct 2016 16:39:42 +0100 Subject: [PATCH 4/8] Fix branch name --- .travis.yml | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/.travis.yml b/.travis.yml index eced1a8..b25ffcf 100644 --- a/.travis.yml +++ b/.travis.yml @@ -30,7 +30,7 @@ perl: - "5.22" before_script: - - git clone --depth 1 --single-branch --branch dev https://github.com/cancerit/cgpBigWig.git + - git clone --depth 1 --single-branch --branch develop https://github.com/cancerit/cgpBigWig.git - cd cgpBigWig - ./setup.sh $HOME/wtsi-opt - cd ../ From b8f8eeb233b521e13358e8d85b4b3ea29a19a773 Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Mon, 3 Oct 2016 20:44:31 +0100 Subject: [PATCH 5/8] Remove use of autodie --- lib/PCAP.pm | 11 +++++++++-- 1 file changed, 9 insertions(+), 2 deletions(-) diff --git a/lib/PCAP.pm b/lib/PCAP.pm index 730103b..ada40b4 100644 --- a/lib/PCAP.pm +++ b/lib/PCAP.pm @@ -25,7 +25,7 @@ use Const::Fast qw(const); use base 'Exporter'; use FindBin qw($Bin); use File::Which qw(which); -use autodie qw(:all); +# don't use autodie, only core perl in here our $VERSION = '3.1.0'; our @EXPORT = qw($VERSION _which); @@ -110,11 +110,12 @@ sub _which { sub ref_lengths { my $fai_file = shift; my %ctg_lengths; - open my $FAI, '<', $fai_file; + open my $FAI, '<', $fai_file or die $!; while(my $l = <$FAI>) { my ($ctg, $len) = split /\t/, $l; $ctg_lengths{$ctg} = $len; } + close $FAI; return \%ctg_lengths; } @@ -142,4 +143,10 @@ Output the brief license text for use in help messages. Return the list of tools that should be installed by setup.sh when upgrading from a previous version. +=item ref_lengths + + my $ref_lengths = PCAP::ref_lengths($fai_file); + +Return a hash ref of reference sequence lengths keyed by sequence/contig name. + =back From 385823d62332d6da6078f85e4298a626369c254d Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Thu, 13 Oct 2016 12:47:14 +0100 Subject: [PATCH 6/8] Remove old varaiable --- setup.sh | 2 -- 1 file changed, 2 deletions(-) diff --git a/setup.sh b/setup.sh index a1ef13b..49a0be5 100755 --- a/setup.sh +++ b/setup.sh @@ -17,8 +17,6 @@ SOURCE_BIGFILE="http://www.cpan.org/authors/id/L/LD/LDS/Bio-BigFile-1.07.tar.gz" # for biobambam SOURCE_BBB_BIN_DIST="https://github.com/gt1/biobambam2/releases/download/2.0.54-release-20160802163650/biobambam2-2.0.54-release-20160802163650-x86_64-etch-linux-gnu.tar.gz" -BIODBHTS_INSTALL="https://raw.githubusercontent.com/Ensembl/Bio-HTS/master/INSTALL.pl" - get_distro () { EXT="" if [[ $2 == *.tar.bz2* ]] ; then From 0127a813da6e6e80774c246d918ced681d6cab85 Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Thu, 13 Oct 2016 12:47:27 +0100 Subject: [PATCH 7/8] Update version docs --- CHANGES.md | 18 ++++++++++++++++++ 1 file changed, 18 insertions(+) diff --git a/CHANGES.md b/CHANGES.md index cb603cf..6a84241 100644 --- a/CHANGES.md +++ b/CHANGES.md @@ -1,3 +1,21 @@ +### 3.2.0 +* Move from legacy kent bigwig manipulation code and to [`cgpBigWig`](https://github.com/cancerit/cgpBigWig) + * Faster and handles the huge number of contigs in many new reference builds. + * Resulting changes to underlying installed tools is `bwcat` now `bwjoin` to be more descriptive of actual function. + +### 3.1.1 +Handle recent changes to BioPerl structure + +### 3.1.0 +* Use BWA default for `-T` previously hard coded to `-T 0`. + * Can be passed through `bwa_mem.pl` other ags to bwa via the `-b` option. +* Fix `bam2bedgraph` compilation since changes to underlying libraries +* `bamToBw.pl` - expose read flag filters +* Drop dependancy on `Bio::DB::HTS INSTALL.pl` as can't fix to known good version. +* Added travis CI +* Add support for output directly to CRAM +* `bwa` version upgraded to [0.7.15](https://github.com/lh3/bwa/releases/tag/v0.7.15) + ### 3.0.0 * Threading module now converts currently running step to bash script for following reasons: * Changes logging to use file redirects instead of Capture::Tiny - prevent log bleed into wrong files From bd4700a501e582144fff319088775c06cae8f26d Mon Sep 17 00:00:00 2001 From: Keiran Raine Date: Thu, 13 Oct 2016 14:48:17 +0100 Subject: [PATCH 8/8] Finalised release --- MANIFEST | 2 -- docs.tar.gz | Bin 172292 -> 171600 bytes lib/PCAP.pm | 50 ++------------------------------------------------ 3 files changed, 2 insertions(+), 50 deletions(-) diff --git a/MANIFEST b/MANIFEST index a1775d1..bec8c9e 100644 --- a/MANIFEST +++ b/MANIFEST @@ -10,7 +10,6 @@ bin/gnos_pull.pl bin/monitor.pl bin/xam_coverage_bins.pl bin/xml_to_bas.pl 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$VERSION = '3.2.0'; our @EXPORT = qw($VERSION _which); const my $LICENSE => @@ -38,53 +38,7 @@ const my $LICENSE => #################"; const my $DEFAULT_PATH => 'biobambam,samtools,bwa'; -const my %UPGRADE_PATH => ( # all earlier versions need full upgrade - '1.2.0' => 'biobambam,samtools,bwa', # if later versions have new versions then all preceding need that tool listing - '1.2.1' => 'biobambam,samtools,bwa', - '1.2.2' => 'biobambam,samtools,bwa', - '1.3.0' => 'biobambam,samtools,bwa', - '1.4.0' => 'biobambam,samtools,bwa', - '1.5.0' => 'biobambam,samtools,bwa', - '1.5.1' => 'biobambam,samtools,bwa', - '1.5.2' => 'biobambam,samtools,bwa', - '1.5.3' => 'biobambam,samtools,bwa', - '1.5.4' => 'biobambam,samtools,bwa', - '1.6.0' => 'biobambam,samtools,bwa', - '1.6.1' => 'biobambam,samtools,bwa', - '1.6.2' => 'biobambam,samtools,bwa', - '1.6.3' => 'biobambam,samtools,bwa', - '1.7.0' => 'biobambam,samtools,bwa', - '1.7.1' => 'biobambam,samtools,bwa', - '1.8.0' => 'biobambam,samtools', - '1.8.1' => 'biobambam,samtools', - '1.8.2' => 'biobambam,samtools', - '1.9.0' => 'biobambam,samtools', - '1.9.1' => 'biobambam,samtools', - '1.9.4' => 'biobambam,samtools', - '1.10.0' => 'biobambam,samtools', - '1.11.0' => 'biobambam,samtools', - '1.11.1' => 'biobambam,samtools', - '1.12.0' => 'biobambam,samtools', - '1.12.1' => 'biobambam,samtools', - '1.12.2' => 'biobambam,samtools', - '1.12.3' => 'biobambam,samtools', - '1.13.0' => 'biobambam,samtools', - '1.13.1' => 'biobambam,samtools', - '1.13.2' => 'biobambam,samtools', - '1.14.0' => 'biobambam,samtools', - '2.0.0' => 'biobambam', - '2.0.1' => 'biobambam,samtools', - '2.1.0' => 'biobambam,samtools', - '2.1.2' => 'biobambam,samtools', - '2.1.3' => 'biobambam,samtools', - '2.2.0' => 'biobambam,samtools', - '2.2.1' => 'biobambam', - '2.3.0' => 'biobambam', - '2.4.0' => 'biobambam', - '2.5.0' => 'biobambam', - '3.0.0' => '', - '3.1.0' => '', - '3.1.1' => '', +const my %UPGRADE_PATH => ( # just always install, it's safer ); sub license {