-
Notifications
You must be signed in to change notification settings - Fork 240
/
gff.c
1116 lines (1007 loc) · 43 KB
/
gff.c
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
208
209
210
211
212
213
214
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
254
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
271
272
273
274
275
276
277
278
279
280
281
282
283
284
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
303
304
305
306
307
308
309
310
311
312
313
314
315
316
317
318
319
320
321
322
323
324
325
326
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
395
396
397
398
399
400
401
402
403
404
405
406
407
408
409
410
411
412
413
414
415
416
417
418
419
420
421
422
423
424
425
426
427
428
429
430
431
432
433
434
435
436
437
438
439
440
441
442
443
444
445
446
447
448
449
450
451
452
453
454
455
456
457
458
459
460
461
462
463
464
465
466
467
468
469
470
471
472
473
474
475
476
477
478
479
480
481
482
483
484
485
486
487
488
489
490
491
492
493
494
495
496
497
498
499
500
501
502
503
504
505
506
507
508
509
510
511
512
513
514
515
516
517
518
519
520
521
522
523
524
525
526
527
528
529
530
531
532
533
534
535
536
537
538
539
540
541
542
543
544
545
546
547
548
549
550
551
552
553
554
555
556
557
558
559
560
561
562
563
564
565
566
567
568
569
570
571
572
573
574
575
576
577
578
579
580
581
582
583
584
585
586
587
588
589
590
591
592
593
594
595
596
597
598
599
600
601
602
603
604
605
606
607
608
609
610
611
612
613
614
615
616
617
618
619
620
621
622
623
624
625
626
627
628
629
630
631
632
633
634
635
636
637
638
639
640
641
642
643
644
645
646
647
648
649
650
651
652
653
654
655
656
657
658
659
660
661
662
663
664
665
666
667
668
669
670
671
672
673
674
675
676
677
678
679
680
681
682
683
684
685
686
687
688
689
690
691
692
693
694
695
696
697
698
699
700
701
702
703
704
705
706
707
708
709
710
711
712
713
714
715
716
717
718
719
720
721
722
723
724
725
726
727
728
729
730
731
732
733
734
735
736
737
738
739
740
741
742
743
744
745
746
747
748
749
750
751
752
753
754
755
756
757
758
759
760
761
762
763
764
765
766
767
768
769
770
771
772
773
774
775
776
777
778
779
780
781
782
783
784
785
786
787
788
789
790
791
792
793
794
795
796
797
798
799
800
801
802
803
804
805
806
807
808
809
810
811
812
813
814
815
816
817
818
819
820
821
822
823
824
825
826
827
828
829
830
831
832
833
834
835
836
837
838
839
840
841
842
843
844
845
846
847
848
849
850
851
852
853
854
855
856
857
858
859
860
861
862
863
864
865
866
867
868
869
870
871
872
873
874
875
876
877
878
879
880
881
882
883
884
885
886
887
888
889
890
891
892
893
894
895
896
897
898
899
900
901
902
903
904
905
906
907
908
909
910
911
912
913
914
915
916
917
918
919
920
921
922
923
924
925
926
927
928
929
930
931
932
933
934
935
936
937
938
939
940
941
942
943
944
945
946
947
948
949
950
951
952
953
954
955
956
957
958
959
960
961
962
963
964
965
966
967
968
969
970
971
972
973
974
975
976
977
978
979
980
981
982
983
984
985
986
987
988
989
990
991
992
993
994
995
996
997
998
999
1000
/* The MIT License
Copyright (c) 2023 Genome Research Ltd.
Author: Petr Danecek <[email protected]>
Permission is hereby granted, free of charge, to any person obtaining a copy
of this software and associated documentation files (the "Software"), to deal
in the Software without restriction, including without limitation the rights
to use, copy, modify, merge, publish, distribute, sublicense, and/or sell
copies of the Software, and to permit persons to whom the Software is
furnished to do so, subject to the following conditions:
The above copyright notice and this permission notice shall be included in
all copies or substantial portions of the Software.
THE SOFTWARE IS PROVIDED "AS IS", WITHOUT WARRANTY OF ANY KIND, EXPRESS OR
IMPLIED, INCLUDING BUT NOT LIMITED TO THE WARRANTIES OF MERCHANTABILITY,
FITNESS FOR A PARTICULAR PURPOSE AND NONINFRINGEMENT. IN NO EVENT SHALL THE
AUTHORS OR COPYRIGHT HOLDERS BE LIABLE FOR ANY CLAIM, DAMAGES OR OTHER
LIABILITY, WHETHER IN AN ACTION OF CONTRACT, TORT OR OTHERWISE, ARISING FROM,
OUT OF OR IN CONNECTION WITH THE SOFTWARE OR THE USE OR OTHER DEALINGS IN
THE SOFTWARE.
*/
#include <stdarg.h>
#include <stdio.h>
#include <stdlib.h>
#include <assert.h>
#include <inttypes.h>
#include <string.h>
#include <strings.h>
#include <htslib/hts.h>
#include <htslib/khash.h>
#include <htslib/khash_str2int.h>
#include <htslib/kseq.h>
#include <htslib/bgzf.h>
#include <errno.h>
#include "bcftools.h"
#include "regidx.h"
#include "gff.h"
/*
Helper structures, only for initialization
ftr_t
temporary list of all exons, CDS, UTRs
*/
KHASH_MAP_INIT_INT(int2tscript, gf_tscript_t*)
KHASH_MAP_INIT_INT(int2gene, gf_gene_t*)
typedef struct
{
int type; // GF_CDS, GF_EXON, GF_5UTR, GF_3UTR
uint32_t beg;
uint32_t end;
uint32_t trid;
uint32_t strand:2; // STRAND_{REV,FWD,UNK}
uint32_t phase:2; // 0, 1, 2, or 3 for unknown
uint32_t iseq:28;
}
ftr_t;
/*
Mapping from GFF ID string (such as ENST00000450305 or Zm00001d027230_P001)
to integer id. To keep the memory requirements low, the original version
relied on IDs in the form of a string prefix and a numerical id. However,
it turns out that this assumption is not valid for some ensembl GFFs, see
for example Zea_mays.AGPv4.36.gff3.gz
*/
typedef struct
{
void *str2id; // khash_str2int
int nstr, mstr;
char **str; // numeric id to string
}
id_tbl_t;
typedef struct
{
// all exons, CDS, UTRs
ftr_t *ftr;
int nftr, mftr;
// mapping from gene id to gf_gene_t
kh_int2gene_t *gid2gene;
// mapping from transcript id to tscript, for quick CDS anchoring
kh_int2tscript_t *id2tr;
// sequences
void *seq2int; // str2int hash
char **seq;
int nseq, mseq;
// ignored biotypes
void *ignored_biotypes;
id_tbl_t gene_ids; // temporary table for mapping between gene id (eg. Zm00001d027245) and a numeric idx
// pointers to the current partially processed line
char *id, *id_end, *parent, *parent_end, *biotype, *biotype_end,
*chr, *chr_end, *name, *name_end, *type, *type_end;
}
aux_t;
struct gff_t_
{
const char *fname, *dump_fname;
// the main regidx lookups, from chr:beg-end to overlapping features and
// index iterator
regidx_t *idx_cds, *idx_utr, *idx_exon, *idx_tscript;
// temporary structures, deleted after initializtion
aux_t init;
// mapping between transcript id (eg. Zm00001d027245_T001) and a numeric idx
id_tbl_t tscript_ids;
int strip_chr_names, verbosity;
int force; // force run under various conditions. Currently only to skip out-of-phase transcripts
struct {
int unknown_chr,unknown_tscript_biotype,unknown_strand,unknown_phase,duplicate_id;
int unknown_cds_phase,incomplete_cds,wrong_phase,overlapping_cds;
} warned;
};
static const char *gf_strings_noncoding[] =
{
"MT_rRNA", "MT_tRNA", "lincRNA", "miRNA", "misc_RNA", "rRNA", "snRNA", "snoRNA", "processed_transcript",
"antisense", "macro_lncRNA", "ribozyme", "sRNA", "scRNA", "scaRNA", "sense_intronic", "sense_overlapping",
"pseudogene", "processed_pseudogene", "artifact", "IG_pseudogene", "IG_C_pseudogene", "IG_J_pseudogene",
"IG_V_pseudogene", "TR_V_pseudogene", "TR_J_pseudogene", "MT_tRNA_pseudogene", "misc_RNA_pseudogene",
"miRNA_pseudogene", "ribozyme", "retained_intron", "retrotransposed", "Trna_pseudogene", "transcribed_processed_pseudogene",
"transcribed_unprocessed_pseudogene", "transcribed_unitary_pseudogene", "translated_unprocessed_pseudogene",
"translated_processed_pseudogene", "known_ncRNA", "unitary_pseudogene", "unprocessed_pseudogene",
"LRG_gene", "3_prime_overlapping_ncRNA", "disrupted_domain", "vaultRNA", "bidirectional_promoter_lncRNA", "ambiguous_orf",
"lncRNA"
};
static const char *gf_strings_coding[] = { "protein_coding", "polymorphic_pseudogene", "IG_C", "IG_D", "IG_J", "IG_LV", "IG_V", "TR_C", "TR_D", "TR_J", "TR_V", "NMD", "non_stop_decay"};
static const char *gf_strings_special[] = { "CDS", "exon", "3_prime_UTR", "5_prime_UTR" };
int gff_set(gff_t *gff, gff_opt_t key, ...)
{
va_list args;
switch (key)
{
case dump_fname:
va_start(args, key);
gff->dump_fname = va_arg(args,char*);
va_end(args);
return 0;
case force_out_of_phase:
va_start(args, key);
gff->force = va_arg(args,int);
va_end(args);
return 0;
case strip_chr_names:
va_start(args, key);
gff->strip_chr_names = va_arg(args,int);
va_end(args);
return 0;
case verbosity:
va_start(args, key);
gff->verbosity = va_arg(args,int);
va_end(args);
return 0;
default:
error("The key %d is not supported with gff_set\n",key);
}
return 0;
}
void *gff_get(gff_t *gff, gff_opt_t key)
{
switch (key)
{
case idx_cds: return gff->idx_cds;
case idx_utr: return gff->idx_utr;
case idx_exon: return gff->idx_exon;
case idx_tscript: return gff->idx_tscript;
default:
error("The key %d is not supported with gff_get\n",key);
}
return NULL;
}
const char *gff_id2string(gff_t *gff, id_type_t type, int id) // currently only transcript ids
{
return gff->tscript_ids.str[id];
}
const char *gf_type2gff_string(int type)
{
if ( !GF_is_coding(type) )
{
if ( type < (1<<GF_coding_bit) ) return gf_strings_noncoding[type-1];
type &= (1<<(GF_coding_bit+1)) - 1;
return gf_strings_special[type - 1];
}
type &= (1<<GF_coding_bit) - 1;
return gf_strings_coding[type - 1];
}
/*
gff parsing functions
*/
static inline int feature_set_seq(gff_t *gff, char *chr_beg, char *chr_end)
{
aux_t *aux = &gff->init;
char tmp = chr_end[1];
chr_end[1] = 0;
int iseq;
if ( khash_str2int_get(aux->seq2int, chr_beg, &iseq)!=0 )
{
char *new_chr = strdup(chr_beg);
hts_expand(char*, aux->nseq+1, aux->mseq, aux->seq);
aux->seq[aux->nseq] = new_chr;
iseq = khash_str2int_inc(aux->seq2int, aux->seq[aux->nseq]);
aux->nseq++;
assert( aux->nseq < 1<<29 ); // see gf_gene_t.iseq and ftr_t.iseq
}
chr_end[1] = tmp;
return iseq;
}
static inline char *gff_skip(const char *line, char *ss)
{
while ( *ss && *ss!='\t' ) ss++;
if ( !*ss ) error("[%s:%d %s] Could not parse the line: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
return ss+1;
}
static inline void gff_parse_chr(gff_t *gff, const char *line, char **chr_beg, char **chr_end)
{
char *se = (char*) line;
while ( *se && *se!='\t' ) se++;
if ( !*se ) error("[%s:%d %s] Could not parse the line: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
if ( gff->strip_chr_names && !strncasecmp("chr",line,3) ) line += 3;
*chr_beg = (char*) line;
*chr_end = se-1;
}
static inline char *gff_parse_beg_end(const char *line, char *ss, uint32_t *beg, uint32_t *end)
{
char *se = ss;
*beg = strtol(ss, &se, 10) - 1;
if ( ss==se ) error("[%s:%d %s] Could not parse the line:\n\t%s\n\t%s\n",__FILE__,__LINE__,__FUNCTION__,line,ss);
ss = se+1;
*end = strtol(ss, &se, 10) - 1;
if ( ss==se ) error("[%s:%d %s] Could not parse the line: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
return se+1;
}
static void gff_id_init(id_tbl_t *tbl)
{
memset(tbl, 0, sizeof(*tbl));
tbl->str2id = khash_str2int_init();
}
static void gff_id_destroy(id_tbl_t *tbl)
{
khash_str2int_destroy_free(tbl->str2id);
free(tbl->str);
}
static inline int gff_id_register(id_tbl_t *tbl, char *beg, char *end, uint32_t *id_ptr)
{
char tmp = end[1];
end[1] = 0;
int id;
if ( khash_str2int_get(tbl->str2id, beg, &id) < 0 )
{
id = tbl->nstr++;
hts_expand(char*, tbl->nstr, tbl->mstr, tbl->str);
tbl->str[id] = strdup(beg);
khash_str2int_set(tbl->str2id, tbl->str[id], id);
}
end[1] = tmp;
*id_ptr = id;
return 0;
}
static inline int gff_parse_biotype(char *line)
{
if ( !line ) return -1;
switch (*line)
{
case 'p':
if ( !strncmp(line,"protein_coding",14) ) return GF_PROTEIN_CODING;
else if ( !strncmp(line,"pseudogene",10) ) return GF_PSEUDOGENE;
else if ( !strncmp(line,"processed_transcript",20) ) return GF_PROCESSED_TRANSCRIPT;
else if ( !strncmp(line,"processed_pseudogene",20) ) return GF_PROCESSED_PSEUDOGENE;
else if ( !strncmp(line,"polymorphic_pseudogene",22) ) return GF_POLYMORPHIC_PSEUDOGENE;
break;
case 'a':
if ( !strncmp(line,"artifact",8) ) return GF_ARTIFACT;
else if ( !strncmp(line,"antisense",9) ) return GF_ANTISENSE;
else if ( !strncmp(line,"ambiguous_orf",13) ) return GF_AMBIGUOUS_ORF;
break;
case 'I':
if ( !strncmp(line,"IG_pseudogene",13) ) return GF_IG_PSEUDOGENE;
else if ( !strncmp(line,"IG_C_pseudogene",15) ) return GF_IG_C_PSEUDOGENE;
else if ( !strncmp(line,"IG_J_pseudogene",15) ) return GF_IG_J_PSEUDOGENE;
else if ( !strncmp(line,"IG_V_pseudogene",15) ) return GF_IG_V_PSEUDOGENE;
else if ( !strncmp(line,"IG_C",4) ) return GF_IG_C;
else if ( !strncmp(line,"IG_D",4) ) return GF_IG_D;
else if ( !strncmp(line,"IG_J",4) ) return GF_IG_J;
else if ( !strncmp(line,"IG_V",4) ) return GF_IG_V;
else if ( !strncmp(line,"IG_LV",5) ) return GF_IG_LV;
break;
case 'T':
if ( !strncmp(line,"TR_V_pseudogene",15) ) return GF_TR_V_PSEUDOGENE;
else if ( !strncmp(line,"TR_J_pseudogene",15) ) return GF_TR_J_PSEUDOGENE;
else if ( !strncmp(line,"TR_C",4) ) return GF_TR_C;
else if ( !strncmp(line,"TR_D",4) ) return GF_TR_D;
else if ( !strncmp(line,"TR_J",4) ) return GF_TR_J;
else if ( !strncmp(line,"TR_V",4) ) return GF_TR_V;
break;
case 'M':
if ( !strncmp(line,"Mt_tRNA_pseudogene",18) ) return GF_MT_tRNA_PSEUDOGENE;
else if ( !strncasecmp(line,"Mt_tRNA",7) ) return GF_MT_tRNA;
else if ( !strncasecmp(line,"Mt_rRNA",7) ) return GF_MT_tRNA;
else if ( !strncasecmp(line,"MRNA",4) ) return GF_PROTEIN_CODING;
break;
case 'l':
if ( !strncmp(line,"lincRNA",7) ) return GF_lincRNA;
if ( !strncmp(line,"lncRNA",7) ) return GF_lncRNA;
break;
case 'm':
if ( !strncmp(line,"macro_lncRNA",12) ) return GF_macro_lncRNA;
else if ( !strncmp(line,"misc_RNA_pseudogene",19) ) return GF_misc_RNA_PSEUDOGENE;
else if ( !strncmp(line,"miRNA_pseudogene",16) ) return GF_miRNA_PSEUDOGENE;
else if ( !strncmp(line,"miRNA",5) ) return GF_miRNA;
else if ( !strncmp(line,"misc_RNA",8) ) return GF_MISC_RNA;
else if ( !strncasecmp(line,"mRNA",4) ) return GF_PROTEIN_CODING;
break;
case 'r':
if ( !strncmp(line,"rRNA",4) ) return GF_rRNA;
else if ( !strncmp(line,"ribozyme",8) ) return GF_RIBOZYME;
else if ( !strncmp(line,"retained_intron",15) ) return GF_RETAINED_INTRON;
else if ( !strncmp(line,"retrotransposed",15) ) return GF_RETROTRANSPOSED;
break;
case 's':
if ( !strncmp(line,"snRNA",5) ) return GF_snRNA;
else if ( !strncmp(line,"sRNA",4) ) return GF_sRNA;
else if ( !strncmp(line,"scRNA",5) ) return GF_scRNA;
else if ( !strncmp(line,"scaRNA",6) ) return GF_scaRNA;
else if ( !strncmp(line,"snoRNA",6) ) return GF_snoRNA;
else if ( !strncmp(line,"sense_intronic",14) ) return GF_SENSE_INTRONIC;
else if ( !strncmp(line,"sense_overlapping",17) ) return GF_SENSE_OVERLAPPING;
break;
case 't':
if ( !strncmp(line,"tRNA_pseudogene",15) ) return GF_tRNA_PSEUDOGENE;
else if ( !strncmp(line,"transcribed_processed_pseudogene",32) ) return GF_TRANSCRIBED_PROCESSED_PSEUDOGENE;
else if ( !strncmp(line,"transcribed_unprocessed_pseudogene",34) ) return GF_TRANSCRIBED_UNPROCESSED_PSEUDOGENE;
else if ( !strncmp(line,"transcribed_unitary_pseudogene",30) ) return GF_TRANSCRIBED_UNITARY_PSEUDOGENE;
else if ( !strncmp(line,"translated_unprocessed_pseudogene",33) ) return GF_TRANSLATED_UNPROCESSED_PSEUDOGENE;
else if ( !strncmp(line,"translated_processed_pseudogene",31) ) return GF_TRANSLATED_PROCESSED_PSEUDOGENE;
break;
case 'n':
if ( !strncmp(line,"nonsense_mediated_decay",23) ) return GF_NMD;
else if ( !strncmp(line,"non_stop_decay",14) ) return GF_NON_STOP_DECAY;
break;
case 'N':
if ( !strncmp(line,"NMD",3) ) return GF_NMD;
break;
case 'k':
if ( !strncmp(line,"known_ncrna",11) ) return GF_KNOWN_NCRNA;
break;
case 'u':
if ( !strncmp(line,"unitary_pseudogene",18) ) return GF_UNITARY_PSEUDOGENE;
else if ( !strncmp(line,"unprocessed_pseudogene",22) ) return GF_UNPROCESSED_PSEUDOGENE;
break;
case 'L':
if ( !strncmp(line,"LRG_gene",8) ) return GF_LRG_GENE;
break;
case '3':
if ( !strncasecmp(line,"3prime_overlapping_ncRNA",24) ) return GF_3PRIME_OVERLAPPING_ncRNA;
else if ( !strncasecmp(line,"3_prime_overlapping_ncRNA",25) ) return GF_3PRIME_OVERLAPPING_ncRNA;
break;
case 'd':
if ( !strncmp(line,"disrupted_domain",16) ) return GF_DISRUPTED_DOMAIN;
break;
case 'v':
if ( !strncmp(line,"vaultRNA",8) ) return GF_vaultRNA;
break;
case 'b':
if ( !strncmp(line,"bidirectional_promoter_lncRNA",29) ) return GF_BIDIRECTIONAL_PROMOTER_lncRNA;
break;
}
return 0;
}
static inline int gff_ignored_biotype(gff_t *gff, char *ss, char *se)
{
if ( !ss ) return 0;
char tmp = se[1];
se[1] = 0;
char *key = ss;
int n = 0;
if ( khash_str2int_get(gff->init.ignored_biotypes, ss, &n)!=0 ) key = strdup(ss);
khash_str2int_set(gff->init.ignored_biotypes, key, n+1);
se[1] = tmp;
return 1;
}
static gf_gene_t *gene_init(aux_t *aux, uint32_t gene_id)
{
khint_t k = kh_get(int2gene, aux->gid2gene, (int)gene_id);
gf_gene_t *gene = (k == kh_end(aux->gid2gene)) ? NULL : kh_val(aux->gid2gene, k);
if ( !gene )
{
gene = (gf_gene_t*) calloc(1,sizeof(gf_gene_t));
int ret;
k = kh_put(int2gene, aux->gid2gene, (int)gene_id, &ret);
kh_val(aux->gid2gene,k) = gene;
}
return gene;
}
static void gff_parse_transcript(gff_t *gff, const char *line, ftr_t *ftr)
{
aux_t *aux = &gff->init;
ftr->type = gff_parse_biotype(aux->biotype);
if ( ftr->type <= 0 )
{
char tmp = aux->type_end[1];
aux->type_end[1] = 0;
ftr->type = gff_parse_biotype(aux->type);
aux->type_end[1] = tmp;
}
if ( ftr->type <= 0 )
{
if ( !gff_ignored_biotype(gff,aux->biotype,aux->biotype_end) )
{
if ( gff->verbosity > 0 )
{
if ( !gff->warned.unknown_tscript_biotype || gff->verbosity > 1 )
fprintf(stderr,"Warning: Ignoring transcript with unknown biotype .. %s\n", line);
gff->warned.unknown_tscript_biotype++;
}
}
return;
}
if ( !aux->id )
error("[%s:%d %s] Could not parse the line, neither \"ID=transcript:\" nor \"ID=\" substring is present: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
if ( !aux->parent )
error("[%s:%d %s] Could not parse the line, neither \"Parent=gene:\" nor \"Parent=\" substring is present: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
uint32_t trid,gene_id;
gff_id_register(&gff->tscript_ids, aux->id, aux->id_end, &trid);
gff_id_register(&aux->gene_ids, aux->parent, aux->parent_end, &gene_id);
gf_tscript_t *tr = (gf_tscript_t*) calloc(1,sizeof(gf_tscript_t));
tr->id = trid;
tr->strand = ftr->strand;
tr->gene = gene_init(aux, gene_id);
tr->type = ftr->type;
tr->beg = ftr->beg;
tr->end = ftr->end;
khint_t k;
int ret;
k = kh_put(int2tscript, aux->id2tr, (int)trid, &ret);
kh_val(aux->id2tr,k) = tr;
}
// register exon, CDS, UTR
static void gff_parse_exon(gff_t *gff, const char *line, ftr_t *ftr)
{
aux_t *aux = &gff->init;
if ( !aux->parent )
error("[%s:%d %s] Could not parse the line, neither \"Parent=transcript:\" nor \"Parent=\" substring found: %s\n",__FILE__,__LINE__,__FUNCTION__,line);
// associate with transcript id
gff_id_register(&gff->tscript_ids, aux->parent, aux->parent_end, &ftr->trid);
if ( ftr->strand==STRAND_UNK && gff->verbosity > 0 )
{
if ( !gff->warned.unknown_strand || gff->verbosity > 1 )
fprintf(stderr,"Warning: Ignoring GFF feature with unknown strand .. %s\n",line);
gff->warned.unknown_strand++;
}
if ( ftr->phase==CDS_PHASE_UNKN && gff->verbosity > 0 )
{
if ( !gff->warned.unknown_phase|| gff->verbosity > 1 )
fprintf(stderr,"Warning: Ignoring GFF feature with unknown phase .. %s\n",line);
gff->warned.unknown_phase++;
}
ftr->iseq = feature_set_seq(gff, aux->chr,aux->chr_end);
}
static void gff_parse_gene(gff_t *gff, const char *line, ftr_t *ftr)
{
aux_t *aux = &gff->init;
if ( !aux->id ) return;
uint32_t gene_id;
gff_id_register(&aux->gene_ids, aux->id, aux->id_end, &gene_id);
gf_gene_t *gene = gene_init(aux, gene_id);
if ( gene->name )
{
if ( !gff->warned.duplicate_id || gff->verbosity > 1 )
fprintf(stderr,"Warning: The GFF contains features with duplicate id .. %s\n",line);
gff->warned.duplicate_id++;
return;
}
gene->iseq = feature_set_seq(gff, aux->chr,aux->chr_end);
gene->beg = ftr->beg;
gene->end = ftr->end;
gene->strand = ftr->strand;
gene->id = gene_id;
if ( aux->name )
{
gene->name = (char*) malloc(aux->name_end - aux->name + 2);
memcpy(gene->name,aux->name,aux->name_end - aux->name + 1);
gene->name[aux->name_end - aux->name + 1] = 0;
}
else
gene->name = strdup(aux->gene_ids.str[gene_id]); // Name=<GeneName> field is not present, use the gene ID instead
}
// Returns 0 for exons,CDS,UTRs to indicate these need to be pruned later and regidx built on them,
// or -1 to indicate the structure needs not be saved (either because of an error or because saved
// as transcript or gene.)
static int gff_parse_line(gff_t *gff, char *line, ftr_t *ftr)
{
// - skip empty lines and commented lines
// - columns
// 1. chr
// 2. <skip>
// 3. CDS, transcript, gene, ...
// 4-5. beg,end
// 6. <skip>
// 7. strand
// 8. phase
// 9. Parent=transcript:ENST(\d+);ID=...;biotype=... etc
char *ss = line;
if ( !*ss ) return -1; // skip blank lines
if ( *ss=='#' ) return -1; // skip comments
aux_t *aux = &gff->init;
gff_parse_chr(gff, line, &aux->chr, &aux->chr_end);
ss = gff_skip(line, aux->chr_end + 2);
// 3rd column: is this a CDS, transcript, gene, etc.. The parsing order by frequency in Homo_sapiens.GRCh37.87.gff3
int is_gene_line = 0;
ftr->type = 0;
aux->type = ss;
if ( !strncmp("exon\t",ss,5) ) { ftr->type = GF_EXON; ss += 5; }
else if ( !strncmp("CDS\t",ss,4) ) { ftr->type = GF_CDS; ss += 4; }
else if ( !strncmp("three_prime_UTR\t",ss,16) ) { ftr->type = GF_UTR3; ss += 16; }
else if ( !strncmp("five_prime_UTR\t",ss,15) ) { ftr->type = GF_UTR5; ss += 15; }
else if ( !strncmp("biological_region\t",ss,18) ) { return -1; } // skip
else if ( !strncmp("gene\t",ss,5) ) { is_gene_line = 1; ss += 5; }
else ss = gff_skip(line, ss);
aux->type_end = ss - 1;
// 4-5th columns: beg,end
ss = gff_parse_beg_end(line, ss, &ftr->beg,&ftr->end);
// 6th column: skip
ss = gff_skip(line, ss);
// 7th column: strand
ftr->strand = -1;
if ( *ss == '+' ) ftr->strand = STRAND_FWD;
else if ( *ss == '-' ) ftr->strand = STRAND_REV;
else ftr->strand = STRAND_UNK;
ss += 2;
// 8th column: phase (codon offset)
ftr->phase = CDS_PHASE_UNKN;
if ( *ss == '0' ) ftr->phase = 0;
else if ( *ss == '1' ) ftr->phase = 1;
else if ( *ss == '2' ) ftr->phase = 2;
else if ( *ss == '.' ) ftr->phase = CDS_PHASE_UNKN; // exons and even CDS in some GFFs do not have phase
ss += 2;
// 9th column: id, parent, name, biotype
aux->name = NULL, aux->id = NULL, aux->parent = NULL, aux->biotype = NULL;
while ( *ss )
{
char *es = ss;
while ( *es && *es!=';' ) es++;
if ( !strncmp(ss,"ID=",3) )
{
ss += 3;
aux->id_end = es - 1;
aux->id = ss;
if ( !strncmp(ss,"gene:",5) ) { aux->id += 5; is_gene_line = 1; }
else if ( !strncmp(ss,"transcript:",11) ) aux->id += 11;
}
else if ( !strncmp(ss,"Name=",5) ) { aux->name = ss + 5; aux->name_end = es - 1; }
else if ( !strncmp(ss,"Parent=",7) )
{
ss += 7;
aux->parent_end = es - 1;
aux->parent = ss;
if ( !strncmp(ss,"gene:",5) ) aux->parent += 5;
else if ( !strncmp(ss,"transcript:",11) ) aux->parent += 11;
}
else if ( !strncmp(ss,"biotype=",8) ) { aux->biotype = ss + 8; aux->biotype_end = es - 1; }
else if ( !strncmp(ss,"gene_biotype=",13) ) { aux->biotype = ss + 13; aux->biotype_end = es - 1; }
if ( !*es ) break;
ss = es + 1;
}
if ( is_gene_line || !aux->parent )
{
gff_parse_gene(gff, line, ftr);
return -1;
}
if ( ftr->type )
{
gff_parse_exon(gff, line, ftr);
return 0;
}
gff_parse_transcript(gff, line, ftr);
return -1;
}
static int cmp_cds_ptr(const void *a, const void *b)
{
// comparison function for qsort of transcripts's CDS
if ( (*((gf_cds_t**)a))->beg < (*((gf_cds_t**)b))->beg ) return -1;
if ( (*((gf_cds_t**)a))->beg > (*((gf_cds_t**)b))->beg ) return 1;
return 0;
}
static inline void chr_beg_end(aux_t *aux, int iseq, char **chr_beg, char **chr_end)
{
*chr_beg = *chr_end = aux->seq[iseq];
while ( (*chr_end)[1] ) (*chr_end)++;
}
static gf_tscript_t *tscript_init(aux_t *aux, uint32_t trid)
{
khint_t k = kh_get(int2tscript, aux->id2tr, (int)trid);
gf_tscript_t *tr = (k == kh_end(aux->id2tr)) ? NULL : kh_val(aux->id2tr, k);
assert( tr );
return tr;
}
static void register_cds(gff_t *gff, ftr_t *ftr)
{
// Make the CDS searchable via idx_cds. Note we do not malloc tr->cds just yet.
// ftr is the result of parsing a gff CDS line
aux_t *aux = &gff->init;
gf_tscript_t *tr = tscript_init(aux, ftr->trid);
if ( tr->strand != ftr->strand ) error("Conflicting strand in transcript %"PRIu32" .. %d vs %d\n",ftr->trid,tr->strand,ftr->strand);
gf_cds_t *cds = (gf_cds_t*) malloc(sizeof(gf_cds_t));
cds->tr = tr;
cds->beg = ftr->beg;
cds->len = ftr->end - ftr->beg + 1;
cds->icds = 0; // to keep valgrind on mac happy
cds->phase = ftr->phase;
hts_expand(gf_cds_t*,tr->ncds+1,tr->mcds,tr->cds);
tr->cds[tr->ncds++] = cds;
}
static void register_utr(gff_t *gff, ftr_t *ftr)
{
aux_t *aux = &gff->init;
gf_utr_t *utr = (gf_utr_t*) malloc(sizeof(gf_utr_t));
utr->which = ftr->type==GF_UTR3 ? prime3 : prime5;
utr->beg = ftr->beg;
utr->end = ftr->end;
utr->tr = tscript_init(aux, ftr->trid);
char *chr_beg, *chr_end;
chr_beg_end(&gff->init, utr->tr->gene->iseq, &chr_beg, &chr_end);
regidx_push(gff->idx_utr, chr_beg,chr_end, utr->beg,utr->end, &utr);
}
static void register_exon(gff_t *gff, ftr_t *ftr)
{
aux_t *aux = &gff->init;
gf_exon_t *exon = (gf_exon_t*) malloc(sizeof(gf_exon_t));
exon->beg = ftr->beg;
exon->end = ftr->end;
exon->tr = tscript_init(aux, ftr->trid);
char *chr_beg, *chr_end;
chr_beg_end(&gff->init, exon->tr->gene->iseq, &chr_beg, &chr_end);
regidx_push(gff->idx_exon, chr_beg,chr_end, exon->beg - N_SPLICE_REGION_INTRON, exon->end + N_SPLICE_REGION_INTRON, &exon);
}
static void tscript_init_cds(gff_t *gff)
{
aux_t *aux = &gff->init;
// Sort CDS in all transcripts, set offsets, check their phase, length, create index (idx_cds)
khint_t k;
for (k=0; k<kh_end(aux->id2tr); k++)
{
if ( !kh_exist(aux->id2tr, k) ) continue;
gf_tscript_t *tr = (gf_tscript_t*) kh_val(aux->id2tr, k);
// position-to-tscript lookup
char *chr_beg, *chr_end;
chr_beg_end(aux, tr->gene->iseq, &chr_beg, &chr_end);
regidx_push(gff->idx_tscript, chr_beg, chr_end, tr->beg, tr->end, &tr);
if ( !tr->ncds ) continue; // transcript with no CDS
// sort CDs
qsort(tr->cds, tr->ncds, sizeof(gf_cds_t*), cmp_cds_ptr);
// trim non-coding start
int i, len = 0;
if ( tr->strand==STRAND_FWD )
{
if ( tr->cds[0]->phase != CDS_PHASE_UNKN )
{
if ( tr->cds[0]->phase ) tr->trim |= TRIM_5PRIME;
tr->cds[0]->beg += tr->cds[0]->phase;
tr->cds[0]->len -= tr->cds[0]->phase;
tr->cds[0]->phase = 0;
}
// sanity check phase; the phase number in gff tells us how many bases to skip in this
// feature to reach the first base of the next codon
int tscript_ok = 1;
for (i=0; i<tr->ncds; i++)
{
if ( tr->cds[i]->phase == CDS_PHASE_UNKN )
{
if ( gff->verbosity > 0 )
{
if ( !gff->warned.unknown_cds_phase || gff->verbosity > 1 )
fprintf(stderr,"Warning: CDS with unknown phase, could not verify reading frame in transcript %s\n",gff->tscript_ids.str[tr->id]);
gff->warned.unknown_cds_phase++;
}
len += tr->cds[i]->len;
continue;
}
int phase = tr->cds[i]->phase ? 3 - tr->cds[i]->phase : 0;
if ( phase!=len%3 )
{
if ( !gff->force )
error("Error: GFF3 assumption failed for transcript %s, CDS=%"PRIu32": phase!=len%%3 (phase=%d, len=%d). Use the --force option to proceed anyway (at your own risk).\n",
gff->tscript_ids.str[tr->id],tr->cds[i]->beg+1,phase,len);
if ( gff->verbosity > 0 )
{
if ( !gff->warned.wrong_phase || gff->verbosity > 1 )
fprintf(stderr,"Warning: The GFF has inconsistent phase column in transcript %s, skipping. CDS pos=%"PRIu32": phase!=len%%3 (phase=%d, len=%d)\n",
gff->tscript_ids.str[tr->id],tr->cds[i]->beg+1,phase,len);
gff->warned.wrong_phase++;
}
tscript_ok = 0;
break;
}
len += tr->cds[i]->len;
}
if ( !tscript_ok ) continue; // skip this transcript
}
else if ( tr->strand==STRAND_REV )
{
if ( tr->cds[tr->ncds-1]->phase != CDS_PHASE_UNKN )
{
// Check that the phase is not bigger than CDS length. Curiously, this can really happen,
// see Mus_musculus.GRCm38.85.gff3.gz, transcript:ENSMUST00000163141.
// This also fixes phase of 5' incomplete CDS, see test/csq/ENST00000520868/ENST00000520868.gff
// todo: the same for the fwd strand
i = tr->ncds - 1;
int phase = tr->cds[i]->phase;
if ( phase ) tr->trim |= TRIM_5PRIME;
while ( i>=0 && phase > tr->cds[i]->len )
{
phase -= tr->cds[i]->len;
tr->cds[i]->phase = 0;
tr->cds[i]->len = 0;
i--;
}
if ( gff->verbosity > 0 && tr->cds[i]->phase )
{
if ( !gff->warned.incomplete_cds || gff->verbosity > 1 )
fprintf(stderr,"Note: truncated transcript %s with incomplete CDS (this is very common)\n",gff->tscript_ids.str[tr->id]);
gff->warned.incomplete_cds++;
}
tr->cds[i]->len -= tr->cds[i]->phase;
tr->cds[i]->phase = 0;
}
// sanity check phase
int tscript_ok = 1;
for (i=tr->ncds-1; i>=0; i--)
{
if ( tr->cds[i]->phase == CDS_PHASE_UNKN )
{
if ( gff->verbosity > 0 )
{
if ( !gff->warned.unknown_cds_phase || gff->verbosity > 1 )
fprintf(stderr,"Warning: CDS with unknown phase, could not verify reading frame in transcript %s\n",gff->tscript_ids.str[tr->id]);
gff->warned.unknown_cds_phase++;
}
len += tr->cds[i]->len;
continue;
}
int phase = tr->cds[i]->phase ? 3 - tr->cds[i]->phase : 0;
if ( phase!=len%3 )
{
if ( !gff->force )
error("Error: GFF3 assumption failed for transcript %s, CDS=%"PRIu32": phase!=len%%3 (phase=%d, len=%d). Use the --force option to proceed anyway (at your own risk).\n",
gff->tscript_ids.str[tr->id],tr->cds[i]->beg+1,phase,len);
if ( gff->verbosity > 0 )
{
if ( !gff->warned.wrong_phase || gff->verbosity > 1 )
fprintf(stderr,"Warning: The GFF has inconsistent phase column in transcript %s, skipping. CDS pos=%"PRIu32": phase!=len%%3 (phase=%d, len=%d)\n",
gff->tscript_ids.str[tr->id],tr->cds[i]->beg+1,phase,len);
gff->warned.wrong_phase++;
}
tscript_ok = 0;
break;
}
len += tr->cds[i]->len;
}
if ( !tscript_ok ) continue; // skip this transcript
}
else
continue; // unknown strand
// set len. At the same check that CDS within a transcript do not overlap
len = 0;
for (i=0; i<tr->ncds; i++)
{
tr->cds[i]->icds = i;
len += tr->cds[i]->len;
if ( !i ) continue;
gf_cds_t *a = tr->cds[i-1];
gf_cds_t *b = tr->cds[i];
if ( a->beg + a->len - 1 >= b->beg )
{
if ( gff->verbosity > 0 )
{
if ( !gff->warned.overlapping_cds || gff->verbosity > 1 )
fprintf(stderr,"Warning: GFF contains overlapping CDS %s, %"PRIu32"-%"PRIu32" and %"PRIu32"-%"PRIu32" (ribosomal slippage?)\n",
gff->tscript_ids.str[tr->id], a->beg+1,a->beg+a->len, b->beg+1,b->beg+b->len);
gff->warned.overlapping_cds++;
}
}
}
if ( len%3 != 0 )
{
// There are 13k transcripts with incomplete 3' CDS. See for example ENST00000524289
// http://sep2015.archive.ensembl.org/Homo_sapiens/Transcript/Sequence_cDNA?db=core;g=ENSG00000155868;r=5:157138846-157159019;t=ENST00000524289
// Also, the incomplete CDS can be too short (1 or 2bp), so it is not enough to trim the last one.
if ( gff->verbosity > 0 )
{
if ( !gff->warned.incomplete_cds || gff->verbosity > 1 )
fprintf(stderr,"Note: truncated transcript %s with incomplete CDS (this is very common)\n",gff->tscript_ids.str[tr->id]);
gff->warned.incomplete_cds++;
}
tr->trim |= TRIM_3PRIME;
if ( tr->strand==STRAND_FWD )
{
i = tr->ncds - 1;
while ( i>=0 && len%3 )
{
int dlen = tr->cds[i]->len >= len%3 ? len%3 : tr->cds[i]->len;
tr->cds[i]->len -= dlen;
len -= dlen;
i--;
}
}
else if ( tr->strand==STRAND_REV )
{
i = 0;
while ( i<tr->ncds && len%3 )
{
int dlen = tr->cds[i]->len >= len%3 ? len%3 : tr->cds[i]->len;
tr->cds[i]->len -= dlen;
tr->cds[i]->beg += dlen;
len -= dlen;
i++;
}
}
}
// set CDS offsets and insert into regidx
len=0;
for (i=0; i<tr->ncds; i++)
{
tr->cds[i]->pos = len;
len += tr->cds[i]->len;
regidx_push(gff->idx_cds, chr_beg,chr_end, tr->cds[i]->beg,tr->cds[i]->beg+tr->cds[i]->len-1, &tr->cds[i]);
}
}
}
static void regidx_free_gf(void *payload) { free(*((gf_cds_t**)payload)); }
static void regidx_free_tscript(void *payload) { gf_tscript_t *tr = *((gf_tscript_t**)payload); free(tr->cds); free(tr); }
static int gff_dump(gff_t *gff, const char *fname)
{
BGZF *out = bgzf_open(fname,"wg");
if ( !out ) error("Failed to open %s: %s\n", fname, strerror(errno));
kstring_t str = {0,0,0};
khint_t k;
for (k=0; k<kh_end(gff->init.gid2gene); k++)
{
if ( !kh_exist(gff->init.gid2gene, k) ) continue;
gf_gene_t *gene = (gf_gene_t*) kh_val(gff->init.gid2gene, k);
char *gene_id = gff->init.gene_ids.str[gene->id];
str.l = 0;
ksprintf(&str,"%s\t.\tgene\t%"PRIu32"\t%"PRIu32"\t.\t%c\t.\tID=%s;Name=%s;used=%d\n",gff->init.seq[gene->iseq],gene->beg+1,gene->end+1,gene->strand==STRAND_FWD?'+':(gene->strand==STRAND_REV?'-':'.'),gene_id,gene->name,gene->used);
if ( bgzf_write(out, str.s, str.l) != str.l ) error("Error writing %s: %s\n", fname, strerror(errno));
}
regitr_t *itr = regitr_init(gff->idx_tscript);
while ( regitr_loop(itr) )
{
gf_tscript_t *tr = regitr_payload(itr, gf_tscript_t*);
char *gene_id = gff->init.gene_ids.str[tr->gene->id];
const char *type = tr->type==GF_PROTEIN_CODING ? "mRNA" : gf_type2gff_string(tr->type);
str.l = 0;
ksprintf(&str,"%s\t.\t%s\t%"PRIu32"\t%"PRIu32"\t.\t%c\t.\tID=%s;Parent=%s;biotype=%s;used=%d\n",itr->seq,type,itr->beg+1,itr->end+1,tr->strand==STRAND_FWD?'+':(tr->strand==STRAND_REV?'-':'.'),gff->tscript_ids.str[tr->id],gene_id,gf_type2gff_string(tr->type),tr->used);
if ( bgzf_write(out, str.s, str.l) != str.l ) error("Error writing %s: %s\n", fname, strerror(errno));
}
regitr_destroy(itr);
itr = regitr_init(gff->idx_cds);
while ( regitr_loop(itr) )
{
gf_cds_t *cds = regitr_payload(itr,gf_cds_t*);
gf_tscript_t *tr = cds->tr;
str.l = 0;
ksprintf(&str,"%s\t.\tCDS\t%"PRIu32"\t%"PRIu32"\t.\t%c\t%c\tParent=%s\n",itr->seq,cds->beg+1,cds->beg+cds->len,tr->strand==STRAND_FWD?'+':(tr->strand==STRAND_REV?'-':'.'),cds->phase==3?'.':cds->phase+(int)'0',gff->tscript_ids.str[tr->id]);
if ( bgzf_write(out, str.s, str.l) != str.l ) error("Error writing %s: %s\n", fname, strerror(errno));
}
regitr_destroy(itr);
itr = regitr_init(gff->idx_utr);
while ( regitr_loop(itr) )
{
gf_utr_t *utr = regitr_payload(itr,gf_utr_t*);
gf_tscript_t *tr = utr->tr;
str.l = 0;
ksprintf(&str,"%s\t.\t%s_prime_UTR\t%"PRIu32"\t%"PRIu32"\t.\t%c\t.\tParent=%s\n",itr->seq,utr->which==prime3?"three":"five",utr->beg+1,utr->end+1,tr->strand==STRAND_FWD?'+':(tr->strand==STRAND_REV?'-':'.'),gff->tscript_ids.str[tr->id]);
if ( bgzf_write(out, str.s, str.l) != str.l ) error("Error writing %s: %s\n", fname, strerror(errno));
}
regitr_destroy(itr);
itr = regitr_init(gff->idx_exon);
while ( regitr_loop(itr) )
{
gf_exon_t *exon = regitr_payload(itr,gf_exon_t*);
gf_tscript_t *tr = exon->tr;
str.l = 0;
ksprintf(&str,"%s\t.\texon\t%"PRIu32"\t%"PRIu32"\t.\t%c\t.\tParent=%s\n",itr->seq,exon->beg+1,exon->end+1,tr->strand==STRAND_FWD?'+':(tr->strand==STRAND_REV?'-':'.'),gff->tscript_ids.str[tr->id]);
if ( bgzf_write(out, str.s, str.l) != str.l ) error("Error writing %s: %s\n", fname, strerror(errno));
}
regitr_destroy(itr);
if ( bgzf_close(out)!=0 ) error("Error: close failed .. %s\n", fname);
free(str.s);
return 0;
}
int gff_parse(gff_t *gff)
{
if ( gff->verbosity > 0 ) fprintf(stderr,"Parsing %s ...\n", gff->fname);
aux_t *aux = &gff->init;
aux->seq2int = khash_str2int_init(); // chrom's numeric id
aux->gid2gene = kh_init(int2gene); // gene id to gf_gene_t, for idx_gene
aux->id2tr = kh_init(int2tscript); // transcript id to tscript_t
gff->idx_tscript = regidx_init(NULL, NULL, regidx_free_tscript, sizeof(gf_tscript_t*), NULL);
aux->ignored_biotypes = khash_str2int_init();
gff_id_init(&aux->gene_ids);
gff_id_init(&gff->tscript_ids);
// parse gff
kstring_t str = {0,0,0};
htsFile *fp = hts_open(gff->fname,"r");
if ( !fp ) error("Failed to read %s\n", gff->fname);
while ( hts_getline(fp, KS_SEP_LINE, &str) > 0 )
{
hts_expand(ftr_t, aux->nftr+1, aux->mftr, aux->ftr);
int ret = gff_parse_line(gff, str.s, aux->ftr + aux->nftr);
if ( !ret ) aux->nftr++;
}
free(str.s);
if ( hts_close(fp)!=0 ) error("Close failed: %s\n", gff->fname);
// process gff information: connect CDS and exons to transcripts
gff->idx_cds = regidx_init(NULL, NULL, regidx_free_gf, sizeof(gf_cds_t*), NULL);