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| Original file line number | Diff line number | Diff line change |
|---|---|---|
|
|
@@ -19,6 +19,7 @@ | |
| from .parser import parse_ref, parse_bed, check_fasta | ||
| from .helper import logger, map_fusion_to_iso, fix_bed, generate_bed | ||
| from collections import defaultdict | ||
| import tempfile | ||
|
|
||
| __author__ = [ | ||
| 'Xiao-Ou Zhang ([email protected])', | ||
|
|
@@ -30,15 +31,17 @@ | |
|
|
||
| @logger | ||
| def annotate(options): | ||
| # create temporary annotated fusion file | ||
| fusion_tmp = tempfile.TemporaryFile(mode='w+') | ||
| # annotate fusion junctions | ||
| annotate_fusion(options['--ref'], options['--bed'], | ||
| annotate_fusion(options['--ref'], options['--bed'], fusion_tmp, | ||
| secondary_flag=options['--low-confidence']) | ||
| # fix fusion juncrions | ||
| fix_fusion(options['--ref'], options['--genome'], options['--output'], | ||
| fix_fusion(options['--ref'], options['--genome'], fusion_tmp, options['--output'], | ||
| options['--no-fix'], secondary_flag=options['--low-confidence']) | ||
|
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|
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| def annotate_fusion(ref_f, junc_bed, secondary_flag=0, denovo_flag=0): | ||
| def annotate_fusion(ref_f, junc_bed, fusion_tmp, secondary_flag=0, denovo_flag=0): | ||
| """ | ||
| Align fusion juncrions to gene annotations | ||
| """ | ||
|
|
@@ -48,96 +51,96 @@ def annotate_fusion(ref_f, junc_bed, secondary_flag=0, denovo_flag=0): | |
| fusion_bed = junc_bed | ||
| fusions, fusion_index = parse_bed(fusion_bed) # fusion junctions | ||
| total = set() | ||
| annotated_fusion_f = 'annotated_fusion.txt.tmp' | ||
| with open(annotated_fusion_f, 'w') as outf: | ||
| for chrom in chrom_info: | ||
| # overlap gene annotations with fusion juncrions | ||
| result = [] | ||
| # overlap genes | ||
| if chrom in genes: | ||
| result += Interval.overlapwith(genes[chrom].interval, | ||
| fusions[chrom]) | ||
| # overlap novel genes in denovo mode | ||
| if denovo_flag and chrom in novel_genes: | ||
| result += Interval.overlapwith(novel_genes[chrom].interval, | ||
| fusions[chrom]) | ||
| for itl in result: | ||
| # extract gene annotations | ||
| iso = list([x for x in itl[2:] if x.startswith('iso')]) | ||
| # for each overlapped fusion junction | ||
| for fus in itl[(2 + len(iso)):]: | ||
| reads = fus.split()[1] | ||
| fus_start, fus_end = fusion_index[fus] | ||
| fus_loc = '%s\t%d\t%d\tFUSIONJUNC/%s' % (chrom, fus_start, | ||
| fus_end, reads) | ||
| edge_annotations = [] # first or last exon flag | ||
| secondary_exon = defaultdict(dict) # secondary exons | ||
| annotate_flag = 0 | ||
| for iso_id in iso: | ||
| g, i, c, s = iso_id.split()[1:] | ||
| start = gene_info[iso_id][0][0] | ||
| end = gene_info[iso_id][-1][-1] | ||
| # fusion junction excesses boundaries of gene | ||
| # annotation | ||
| if fus_start < start - 10 or fus_end > end + 10: | ||
| if not secondary_flag: | ||
| continue | ||
| (fusion_info, | ||
| index, | ||
| edge, | ||
| secondary) = map_fusion_to_iso(fus_start, | ||
| fus_end, s, | ||
| gene_info[iso_id]) | ||
| if fusion_info: | ||
| annotate_flag += 1 | ||
| bed_info = '\t'.join([fus_loc, '0', s, | ||
| str(fus_start), | ||
| str(fus_start), '0,0,0']) | ||
| bed = '\t'.join([bed_info, fusion_info, g, i, | ||
| index]) | ||
| if not edge: # not first or last exon | ||
| outf.write(bed + '\n') | ||
| total.add(fus) | ||
| else: # first or last exon | ||
| edge_annotations.append(bed) | ||
| elif secondary_flag and secondary is not None: | ||
| li, ri = secondary | ||
| gene = ':'.join([g, s]) | ||
| if li is not None: | ||
| li = str(li) | ||
| secondary_exon['left'][gene] = ':'.join([i, | ||
| li]) | ||
| if ri is not None: | ||
| ri = str(ri) | ||
| secondary_exon['right'][gene] = ':'.join([i, | ||
| ri]) | ||
| if edge_annotations: | ||
| for bed in edge_annotations: | ||
| outf = fusion_tmp | ||
| for chrom in chrom_info: | ||
| # overlap gene annotations with fusion juncrions | ||
| result = [] | ||
| # overlap genes | ||
| if chrom in genes: | ||
| result += Interval.overlapwith(genes[chrom].interval, | ||
| fusions[chrom]) | ||
| # overlap novel genes in denovo mode | ||
| if denovo_flag and chrom in novel_genes: | ||
| result += Interval.overlapwith(novel_genes[chrom].interval, | ||
| fusions[chrom]) | ||
| for itl in result: | ||
| # extract gene annotations | ||
| iso = list([x for x in itl[2:] if x.startswith('iso')]) | ||
| # for each overlapped fusion junction | ||
| for fus in itl[(2 + len(iso)):]: | ||
| reads = fus.split()[1] | ||
| fus_start, fus_end = fusion_index[fus] | ||
| fus_loc = '%s\t%d\t%d\tFUSIONJUNC/%s' % (chrom, fus_start, | ||
| fus_end, reads) | ||
| edge_annotations = [] # first or last exon flag | ||
| secondary_exon = defaultdict(dict) # secondary exons | ||
| annotate_flag = 0 | ||
| for iso_id in iso: | ||
| g, i, c, s = iso_id.split()[1:] | ||
| start = gene_info[iso_id][0][0] | ||
| end = gene_info[iso_id][-1][-1] | ||
| # fusion junction excesses boundaries of gene | ||
| # annotation | ||
| if fus_start < start - 10 or fus_end > end + 10: | ||
| if not secondary_flag: | ||
| continue | ||
| (fusion_info, | ||
| index, | ||
| edge, | ||
| secondary) = map_fusion_to_iso(fus_start, | ||
| fus_end, s, | ||
| gene_info[iso_id]) | ||
| if fusion_info: | ||
| annotate_flag += 1 | ||
| bed_info = '\t'.join([fus_loc, '0', s, | ||
| str(fus_start), | ||
| str(fus_start), '0,0,0']) | ||
| bed = '\t'.join([bed_info, fusion_info, g, i, | ||
| index]) | ||
| if not edge: # not first or last exon | ||
| outf.write(bed + '\n') | ||
| total.add(fus) | ||
| if secondary_flag and not annotate_flag: | ||
| for gene in secondary_exon['left']: | ||
| if gene in secondary_exon['right']: | ||
| left = secondary_exon['left'][gene] | ||
| right = secondary_exon['right'][gene] | ||
| g, s = gene.split(':') | ||
| # for avoid dup, use fus_loc_new | ||
| fus_loc_new = fus_loc + '\t0\t%s' % s | ||
| outf.write('%s\t%s:%s\t%s:%s\n' % (fus_loc_new, | ||
| g, left, g, | ||
| right)) | ||
| total.add(fus) | ||
| else: # first or last exon | ||
| edge_annotations.append(bed) | ||
| elif secondary_flag and secondary is not None: | ||
| li, ri = secondary | ||
| gene = ':'.join([g, s]) | ||
| if li is not None: | ||
| li = str(li) | ||
| secondary_exon['left'][gene] = ':'.join([i, | ||
| li]) | ||
| if ri is not None: | ||
| ri = str(ri) | ||
| secondary_exon['right'][gene] = ':'.join([i, | ||
| ri]) | ||
| if edge_annotations: | ||
| for bed in edge_annotations: | ||
| outf.write(bed + '\n') | ||
| total.add(fus) | ||
| if secondary_flag and not annotate_flag: | ||
| for gene in secondary_exon['left']: | ||
| if gene in secondary_exon['right']: | ||
| left = secondary_exon['left'][gene] | ||
| right = secondary_exon['right'][gene] | ||
| g, s = gene.split(':') | ||
| # for avoid dup, use fus_loc_new | ||
| fus_loc_new = fus_loc + '\t0\t%s' % s | ||
| outf.write('%s\t%s:%s\t%s:%s\n' % (fus_loc_new, | ||
| g, left, g, | ||
| right)) | ||
| print('Annotated %d fusion junctions!' % len(total)) | ||
|
|
||
|
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||
| def fix_fusion(ref_f, genome_fa, out_file, no_fix, secondary_flag=0, | ||
| def fix_fusion(ref_f, genome_fa, fusion_tmp, out_file, no_fix, secondary_flag=0, | ||
| denovo_flag=0): | ||
| """ | ||
| Realign fusion juncrions | ||
| """ | ||
| print('Start to fix fusion junctions...') | ||
| fa = check_fasta(genome_fa) | ||
| ref = parse_ref(ref_f, 2) | ||
| annotated_fusion_f = 'annotated_fusion.txt.tmp' | ||
| annotated_fusion_f = fusion_tmp | ||
| annotated_fusion_f.seek(0) ## from the start | ||
| fusions, fusion_names, fixed_flag = fix_bed(annotated_fusion_f, ref, fa, | ||
| no_fix, denovo_flag) | ||
| total = 0 | ||
|
|
@@ -216,6 +219,6 @@ def fix_fusion(ref_f, genome_fa, out_file, no_fix, secondary_flag=0, | |
| if secondary_flag: | ||
| secondary_f.close() | ||
|
|
||
| os.remove('annotated_fusion.txt.tmp') | ||
| fusion_tmp.close() | ||
|
|
||
| print('Fixed %d fusion junctions!' % total) | ||
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| Original file line number | Diff line number | Diff line change |
|---|---|---|
|
|
@@ -32,6 +32,7 @@ | |
| import pysam | ||
| from scipy.stats import fisher_exact, binom | ||
| from .genomic_interval import Interval | ||
| import tempfile | ||
|
|
||
| __author__ = [ | ||
| 'Xiao-Ou Zhang ([email protected])', | ||
|
|
@@ -66,11 +67,13 @@ def denovo(options): | |
| print('Warning: no cufflinks directory %s!' % options['--cuff']) | ||
| print('Please run CIRCexplorer2 assembly before this step!') | ||
| ref_path = options['--ref'] | ||
| # create temporary annotated fusion file | ||
| fusion_tmp = tempfile.TemporaryFile(mode='w+') | ||
| # annotate fusion junctions | ||
| annotate_fusion(ref_path, options['--bed'], denovo_flag=1) | ||
| annotate_fusion(ref_path, options['--bed'], fusion_tmp, denovo_flag=1) | ||
| # fix fusion juncrions | ||
| out_f = '%s/circularRNA_full.txt' % denovo_dir | ||
| fix_fusion(ref_path, options['--genome'], out_f, | ||
| fix_fusion(ref_path, options['--genome'], fusion_tmp, out_f, | ||
| options['--no-fix'], denovo_flag=1) | ||
| # extract novel circRNAs | ||
| extract_novel_circ(denovo_dir, options['--ref']) | ||
|
|
||
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