int main(int argc, char *argv[]) {
int check_start = 0, check_stop = 0, check_splice = 0, check_nonsense = 0,
offset5 = 0, offset3 = 0, opt_idx, i, j, indel_strict = 0, no_output = 0,
check_alignment = 0, splice_strict = 0;
int ncons_tested, nkept, nconserved_exons;
int nce_gap_type[NGAP_TYPES], nconsid[NTYPES], nfail[NTYPES];
double Nfrac = 0.05;
char c;
MSA *msa;
GFF_Set *gff;
msa_format_type msa_format = UNKNOWN_FORMAT;
List *keepers, *problems = lst_new_ptr(10),
*ends_adjusted = lst_new_ptr(1), *starts_adjusted = lst_new_ptr(1),
*discards=NULL, *intron_splice = lst_new_ptr(10);
char *rseq_fname = NULL;
FILE *logf = NULL, *mlogf = NULL, *statsf = NULL, *discardf = NULL;
cds_gap_type fshift_mode = FSHIFT_BAD;
char *groupby = "transcript_id";
msa_coord_map *map;
int *countNs, *countCDSs;
FILE *infile;
char *msa_fname;
struct option long_opts[] = {
{"start", 0, 0, 's'},
{"stop", 0, 0, 't'},
{"splice", 0, 0, 'l'},
{"nonsense", 0, 0, 'n'},
{"fshift", 0, 0, 'f'},
{"conserved", 0, 0, 'c'},
{"N-limit", 1, 0, 'N'},
{"clean-gaps", 0, 0, 'e'},
{"indel-strict", 0, 0, 'I'},
{"splice-strict", 0, 0, 'C'},
{"groupby", 1, 0, 'g'},
{"msa-format", 1, 0, 'i'},
{"refseq", 1, 0, 'r'},
{"offset5", 1, 0, 'o'},
{"offset3", 1, 0, 'p'},
{"no-output", 0, 0, 'x'},
{"discards", 1, 0, 'd'},
{"log", 1, 0, 'L'},
{"machine-log", 1, 0, 'M'},
{"stats", 1, 0, 'S'},
{"help", 0, 0, 'h'},
{0, 0, 0, 0}
};
while ((c = (char)getopt_long(argc, argv, "N:i:r:L:M:S:g:d:stlnfceICxh",
long_opts, &opt_idx)) != -1) {
switch(c) {
case 's':
check_alignment = check_start = 1;
break;
case 't':
check_alignment = check_stop = 1;
break;
case 'l':
check_alignment = check_splice = 1;
break;
case 'n':
check_alignment = check_nonsense = 1;
break;
case 'f':
check_alignment = 1;
fshift_mode = FSHIFT_OK;
break;
case 'c':
check_alignment = check_start = check_stop = check_splice = check_nonsense = 1;
if (fshift_mode < FSHIFT_OK) fshift_mode = FSHIFT_OK;
break;
case 'N':
Nfrac = get_arg_dbl_bounds(optarg, 0, 1);
break;
case 'e':
check_alignment = 1;
if (fshift_mode < CLN_GAPS) fshift_mode = CLN_GAPS;
break;
case 'I':
check_alignment = 1;
fshift_mode = NOVRLP_CLN_GAPS;
indel_strict = 1;
break;
case 'C':
check_alignment = check_splice = splice_strict = 1;
break;
case 'g':
groupby = optarg;
break;
case 'i':
msa_format = msa_str_to_format(optarg);
if (msa_format == UNKNOWN_FORMAT) die("Bad alignment format.\n");
break;
case 'r':
rseq_fname = optarg;
break;
case 'o':
offset5 = get_arg_int(optarg);
break;
case 'p':
offset3 = get_arg_int(optarg);
break;
case 'L':
logf = phast_fopen(optarg, "w+");
break;
case 'M':
mlogf = phast_fopen(optarg, "w+");
break;
case 'S':
statsf = phast_fopen(optarg, "w+");
break;
case 'd':
discardf = phast_fopen(optarg, "w+");
break;
case 'x':
no_output = 1;
break;
case 'h':
printf("%s", HELP);
exit(0);
case '?':
die("ERROR: Bad argument. Try the --help option.\n");
}
}
if (optind + 1 >= argc ) {
die("ERROR: Missing required arguments. Try the --help option.\n");
}
set_seed(-1);
gff = gff_read_set(phast_fopen(argv[optind], "r"));
msa_fname = argv[optind+1];
infile = phast_fopen(msa_fname, "r");
if (msa_format == UNKNOWN_FORMAT)
msa_format = msa_format_for_content(infile, 1);
if (msa_format == MAF) {
msa = maf_read(infile,
rseq_fname == NULL ? NULL : phast_fopen(rseq_fname, "r"),
1, NULL, NULL, NULL, -1, TRUE, NULL, NO_STRIP, FALSE);
}
else {
msa = msa_new_from_file_define_format(infile,
msa_format, NULL);
if (msa->ss == NULL)
ss_from_msas(msa, 1, 1, NULL, NULL, NULL, -1, 0);
}
if (!msa->ss->tuple_idx)
die("ERROR: need ordered tuples\n");
msa_remove_N_from_alph(msa); /* for backward compatibility (old SS files) */
if (msa->idx_offset != 0) { /* avoids offset problem */
for (i = 0; i < lst_size(gff->features); i++) {
GFF_Feature *f = lst_get_ptr(gff->features, i);
f->start -= msa->idx_offset;
f->end -= msa->idx_offset;
}
}
/* set up coordinate map; assume GFF is for sequence 1 */
map = msa_build_coord_map(msa, 1);
/* convert all features */
for (i = 0; i < lst_size(gff->features); i++) {
GFF_Feature *f = lst_get_ptr(gff->features, i);
int newstart, newend;
if (f->start < 0 || f->end < f->start)
die("ERROR: bad feature in GFF (start=%d, end=%d).\n",
f->start, f->end);
newstart = msa_map_seq_to_msa(map, f->start);
newend = msa_map_seq_to_msa(map, f->end);
if (newstart < 0 || newend < newstart)
die("ERROR: unable to map coordinates for feature (start=%d, end=%d).\n",
f->start, f->end);
f->start = newstart;
f->end = newend;
}
gff_group(gff, groupby); /* do this after coord conversion, or
group coords and feature coords
will be out of sync */
keepers = lst_new_ptr(lst_size(gff->features));
if (discardf != NULL) discards = lst_new_ptr(lst_size(gff->features));
ncons_tested = nkept = nconserved_exons = 0;
for (i = 0; i < NTYPES; i++) nconsid[i] = 0;
for (i = 0; i < NTYPES; i++) nfail[i] = 0;
for (i = 0; i < NGAP_TYPES; i++) nce_gap_type[i] = 0;
countNs = smalloc(msa->nseqs * sizeof(int));
countCDSs = smalloc(msa->nseqs * sizeof(int));
for (i = 0; i < lst_size(gff->groups); i++) {
GFF_FeatureGroup *group = lst_get_ptr(gff->groups, i);
List *gfeatures = group->features;
GFF_Feature *feat;
status_type status = OKAY;
cds_gap_type gt = FSHIFT_BAD;
problems_clear(problems);
/* make sure have frame info for CDSs */
for (j = 0; j < lst_size(gfeatures); j++) {
feat = lst_get_ptr(gfeatures, j);
if (str_equals_charstr(feat->feature, GFF_CDS_TYPE) &&
feat->frame == GFF_NULL_FRAME)
die("ERROR: Missing frame info for CDS.\n");
}
/* First, exclude stop codons from cds's, if necessary (simplifies
the detection of nonsense mutations). */
exclude_stops(group, starts_adjusted, ends_adjusted);
/* In all cases, discard any group for which the reference sequence
doesn't have valid splice sites or start/stop codons, or has a
premature stop codon */
if (!ref_seq_okay(gfeatures, msa, offset3, indel_strict, splice_strict,
problems)) {
status = BAD_REF;
nfail[BAD_REF]++;
}
else
/* Everything else counts as a potentially valid group */
ncons_tested++;
if (status == OKAY && check_alignment) {
/* only bother with below if
interested in cross-species
conservation */
/* Check first to make sure there's alignment across species in
the cds; if not, there's no need to look at individual
features. */
for (j = 0; j < lst_size(gfeatures); j++) {
feat = lst_get_ptr(gfeatures, j);
if (str_equals_charstr(feat->feature, GFF_CDS_TYPE) &&
is_incomplete_alignment(feat, msa)) {
status = NO_ALN;
nfail[NO_ALN]++;
problem_add(problems, feat, NO_ALN, -1, -1);
break;
}
}
if (status == OKAY) { /* we have alignment and agreement
with the ref seq; now check feature
by feature */
lst_clear(intron_splice);
for (j = 0; j < msa->nseqs; j++) countNs[j] = countCDSs[j] = 0;
for (j = 0; j < lst_size(gfeatures); j++) {
feat = lst_get_ptr(gfeatures, j);
if (feat->end - 1 >= msa->length)
die("ERROR: feature extends beyond alignment (%d >= %d).\n",
feat->end - 1, msa->length);
if (check_start && str_equals_charstr(feat->feature, GFF_START_TYPE)) {
nconsid[BAD_START]++;
if (!is_conserved_start(feat, msa)) {
status = BAD_START;
problem_add(problems, feat, BAD_START, -1, -1);
}
}
else if (check_stop && str_equals_charstr(feat->feature, GFF_STOP_TYPE)) {
nconsid[BAD_STOP]++;
if (!is_conserved_stop(feat, msa)) {
status = BAD_STOP;
problem_add(problems, feat, BAD_STOP, -1, -1);
}
}
else if (check_splice &&
str_equals_charstr(feat->feature, SPLICE_5)) {
nconsid[BAD_5_SPLICE]++;
if (!is_conserved_5splice(feat, msa, offset5, splice_strict)) {
status = BAD_5_SPLICE;
problem_add(problems, feat, BAD_5_SPLICE, -1, -1);
}
else lst_push_ptr(intron_splice, feat);
}
else if (check_splice &&
str_equals_charstr(feat->feature, SPLICE_5_UTR)) {
nconsid[BAD_5_SPLICE_UTR]++;
if (!is_conserved_5splice(feat, msa, offset5, splice_strict)) {
status = BAD_5_SPLICE_UTR;
problem_add(problems, feat, BAD_5_SPLICE_UTR, -1, -1);
}
else lst_push_ptr(intron_splice, feat);
}
else if (check_splice && str_equals_charstr(feat->feature, SPLICE_3)) {
nconsid[BAD_3_SPLICE]++;
if (!is_conserved_3splice(feat, msa, offset3, splice_strict)) {
status = BAD_3_SPLICE;
problem_add(problems, feat, BAD_3_SPLICE, -1, -1);
}
else lst_push_ptr(intron_splice, feat);
}
else if (check_splice && str_equals_charstr(feat->feature, SPLICE_3)) {
nconsid[BAD_3_SPLICE_UTR]++;
if (!is_conserved_3splice(feat, msa, offset3, splice_strict)) {
status = BAD_3_SPLICE_UTR;
problem_add(problems, feat, BAD_3_SPLICE_UTR, -1, -1);
}
else lst_push_ptr(intron_splice, feat);
}
else if (str_equals_charstr(feat->feature, GFF_CDS_TYPE)) {
if (fshift_mode > FSHIFT_BAD
&& (gt = get_cds_gap_type(feat, msa, problems)) < fshift_mode) {
if (status == OKAY || status == NONSENSE) status = FSHIFT;
}
if (check_nonsense && !is_nonsense_clean(feat, msa, problems)) {
if (status == OKAY) status = NONSENSE;
}
if (Nfrac < 1)
get_N_counts(countNs, countCDSs, feat, msa);
}
} /* end loop through features in group */
/* still have to make sure splice sites are paired correctly
(GT-AG, GC-AG, AT-AC) */
if (status == OKAY && !splice_strict && lst_size(intron_splice) >= 2 &&
!are_introns_okay(intron_splice, msa, problems, offset5, offset3))
status = BAD_INTRON;
/* also check fraction of Ns */
if (Nfrac < 1) {
enum {MY_OKAY, MY_FAIL, MY_WARN} Nstatus = MY_OKAY;
for (j = 0; j < msa->nseqs; j++) {
if ((double)countNs[j] / countCDSs[j] > Nfrac) Nstatus = MY_FAIL;
if (Nstatus == MY_OKAY && countNs[j] > 0) Nstatus = MY_WARN;
}
if (Nstatus == MY_FAIL) {
problem_add(problems, NULL, TOO_MANY_Ns, -1, -1);
if (status == OKAY) status = TOO_MANY_Ns;
}
else if (Nstatus == MY_WARN)
problem_add(problems, NULL, WARN_Ns, -1, -1);
}
/* if collecting stats, record counts for failures */
if (statsf != NULL) {
if (status != OKAY) {
for (j = 0; j < lst_size(problems); j++) {
struct Problem *problem = lst_get_ptr(problems, j);
status_type ftype = problem->status;
if ((ftype == FSHIFT || ftype == NONSENSE) &&
status != FSHIFT && status != NONSENSE)
continue; /* don't count secondary frame shifts
and nonsense mutations */
if (ftype == BAD_INTRON && j % 2 == 0)
continue; /* only count one of every pair of these */
nfail[ftype]++;
}
}
/* also keep track of the total number of "conserved exons", and
the number having each kind of gap */
if ((status == OKAY || (status == FSHIFT && gt >= FSHIFT_OK))) {
nconserved_exons++;
nce_gap_type[gt]++; /* number of conserved exons having
given type of gaps */
}
}
} /* end if (status == OKAY) [checks for conserved features] */
} /* end if (status == OKAY && check_alignment) [all cross-species
checks] */
/* now we have looked at the whole group; we just need to do some
final accounting and logging */
if (status == OKAY) {
nkept++;
if (!no_output) {
restore_stops(group, starts_adjusted, ends_adjusted);
for (j = 0; j < lst_size(gfeatures); j++)
lst_push_ptr(keepers, lst_get_ptr(gfeatures, j));
}
if (logf != NULL && lst_size(problems) > 0) /* warnings only */
write_log(logf, group, status, problems, msa, map);
if (mlogf != NULL) {
/* no problem, need to add an okay status to log */
problem_add(problems, NULL, OKAY, -1, -1);
write_machine_log(mlogf, group, problems, map); /* may include
warnings */
}
}
else {
if (discardf != NULL) {
restore_stops(group, starts_adjusted, ends_adjusted);
for (j = 0; j < lst_size(gfeatures); j++)
lst_push_ptr(discards, lst_get_ptr(gfeatures, j));
}
if (logf != NULL)
write_log(logf, group, status, problems, msa, map);
if (mlogf != NULL)
write_machine_log(mlogf, group, problems, map);
}
} /* end loop over groups */
/* write main output and discards */
if (!no_output || discardf != NULL) {
/* first map features back to coord frame of reference seq. */
for (i = 0; i < lst_size(gff->features); i++) {
GFF_Feature *f = lst_get_ptr(gff->features, i);
f->start = msa_map_msa_to_seq(map, f->start) + msa->idx_offset;
f->end = msa_map_msa_to_seq(map, f->end) + msa->idx_offset;
}
if (!no_output) {
gff->features = keepers;
gff_print_set(stdout, gff);
}
if (discardf != NULL) {
gff->features = discards;
gff_print_set(discardf, gff);
}
}
/* dump counts to stats file */
if (statsf != NULL) {
fprintf(statsf, "#%11s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s %12s\n",
"total", "nbad_ref", "nconsid", "nkept", "nno_aln",
"nbad_starts", "(out of)", "nbad_stops", "(out of)",
"nbad_5spl", "(out of)", "nbad_3spl", "(out of)",
"nbad_5utr", "(out of)", "nbad_3utr", "(out of)",
"nbad_intron", "nnons", "nfshifts", "nNs", "ncons_exons",
"nce_ngaps", "nce_nov_cln", "nce_clean", "nce_fshftok");
fprintf(statsf, "%12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d %12d\n",
nfail[BAD_REF]+ncons_tested, nfail[BAD_REF], ncons_tested, nkept,
nfail[NO_ALN], nfail[BAD_START], nconsid[BAD_START],
nfail[BAD_STOP], nconsid[BAD_STOP], nfail[BAD_5_SPLICE],
nconsid[BAD_5_SPLICE], nfail[BAD_3_SPLICE], nconsid[BAD_3_SPLICE],
nfail[BAD_5_SPLICE_UTR], nconsid[BAD_5_SPLICE_UTR],
nfail[BAD_3_SPLICE_UTR], nconsid[BAD_3_SPLICE_UTR],
nfail[BAD_INTRON], nfail[NONSENSE], nfail[FSHIFT],
nfail[TOO_MANY_Ns], nconserved_exons, nce_gap_type[NGAPS],
nce_gap_type[NOVRLP_CLN_GAPS], nce_gap_type[CLN_GAPS],
nce_gap_type[FSHIFT_OK]);
fprintf(statsf, "%s", STATS_DESCRIPTION);
}
if (logf != NULL) phast_fclose(logf);
if (mlogf != NULL) phast_fclose(mlogf);
if (statsf != NULL) phast_fclose(statsf);
if (discardf != NULL) phast_fclose(discardf);
return 0;
}
int main(int argc, char *argv[]) {
char c;
char *msa_fname = NULL;
int opt_idx, i, old_nnodes;
MSA *msa;
List *pruned_names = lst_new_ptr(5), *tmpl;
BDPhyloHmm *bdphmm;
GFF_Set *predictions;
int found = FALSE;
List *ignore_types = lst_new_ptr(1);
struct option long_opts[] = {
{"refseq", 1, 0, 'M'},
{"msa-format", 1, 0, 'i'},
{"refidx", 1, 0, 'r'},
{"rho", 1, 0, 'R'},
{"phi", 1, 0, 'p'},
{"transitions", 1, 0, 't'},
{"expected-length", 1, 0, 'E'},
{"target-coverage", 1, 0, 'C'},
{"seqname", 1, 0, 'N'},
{"idpref", 1, 0, 'P'},
{"indel-model", 1, 0, 'I'},
{"indel-history", 1, 0, 'H'},
{"help", 0, 0, 'h'},
{0, 0, 0, 0}
};
/* arguments and defaults for options */
FILE *refseq_f = NULL, *msa_f = NULL;
msa_format_type msa_format = UNKNOWN_FORMAT;
TreeModel *source_mod;
double rho = DEFAULT_RHO, mu = DEFAULT_MU, nu = DEFAULT_NU,
phi = DEFAULT_PHI, gamma = -1, omega = -1,
alpha_c = -1, beta_c = -1, tau_c = -1,
alpha_n = -1, beta_n = -1, tau_n = -1;
int set_transitions = FALSE, refidx = 1, estim_phi = TRUE,
estim_gamma = TRUE, estim_omega = TRUE;
char *seqname = NULL, *idpref = NULL;
IndelHistory *ih = NULL;
while ((c = getopt_long(argc, argv, "R:t:p:E:C:r:M:i:N:P:I:H:h", long_opts, &opt_idx)) != -1) {
switch (c) {
case 'R':
rho = get_arg_dbl_bounds(optarg, 0, 1);
break;
case 't':
if (optarg[0] != '~') estim_gamma = estim_omega = FALSE;
else optarg = &optarg[1];
set_transitions = TRUE;
tmpl = get_arg_list_dbl(optarg);
if (lst_size(tmpl) != 2)
die("ERROR: bad argument to --transitions.\n");
mu = lst_get_dbl(tmpl, 0);
nu = lst_get_dbl(tmpl, 1);
if (mu <= 0 || mu >= 1 || nu <= 0 || nu >= 1)
die("ERROR: bad argument to --transitions.\n");
lst_free(tmpl);
break;
case 'p':
if (optarg[0] != '~') estim_phi = FALSE;
else optarg = &optarg[1];
phi = get_arg_dbl_bounds(optarg, 0, 1);
break;
case 'E':
if (optarg[0] != '~') estim_omega = FALSE;
else optarg = &optarg[1];
omega = get_arg_dbl_bounds(optarg, 1, INFTY);
mu = 1/omega;
break;
case 'C':
if (optarg[0] != '~') estim_gamma = FALSE;
else optarg = &optarg[1];
gamma = get_arg_dbl_bounds(optarg, 0, 1);
break;
case 'r':
refidx = get_arg_int_bounds(optarg, 0, INFTY);
break;
case 'M':
refseq_f = phast_fopen(optarg, "r");
break;
case 'i':
msa_format = msa_str_to_format(optarg);
if (msa_format == UNKNOWN_FORMAT)
die("ERROR: unrecognized alignment format.\n");
break;
case 'N':
seqname = optarg;
break;
case 'P':
idpref = optarg;
break;
case 'I':
tmpl = get_arg_list_dbl(optarg);
if (lst_size(tmpl) != 3 && lst_size(tmpl) != 6)
die("ERROR: bad argument to --indel-model.\n");
alpha_n = lst_get_dbl(tmpl, 0);
beta_n = lst_get_dbl(tmpl, 1);
tau_n = lst_get_dbl(tmpl, 2);
if (lst_size(tmpl) == 6) {
alpha_c = lst_get_dbl(tmpl, 3);
beta_c = lst_get_dbl(tmpl, 4);
tau_c = lst_get_dbl(tmpl, 5);
}
else {
alpha_c = alpha_n; beta_c = beta_n; tau_c = tau_n;
}
if (alpha_c <= 0 || alpha_c >= 1 || beta_c <= 0 || beta_c >= 1 ||
tau_c <= 0 || tau_c >= 1 || alpha_n <= 0 || alpha_n >= 1 ||
beta_n <= 0 || beta_n >= 1 || tau_n <= 0 || tau_n >= 1)
die("ERROR: bad argument to --indel-model.\n");
break;
case 'H':
fprintf(stderr, "Reading indel history from %s...\n", optarg);
ih = ih_new_from_file(phast_fopen(optarg, "r"));
break;
case 'h':
printf("%s", HELP);
exit(0);
case '?':
die("Bad argument. Try 'dless -h'.\n");
}
}
if (optind != argc - 1)
die("Missing alignment file or model file. Try 'dless -h'.\n");
if (set_transitions && (gamma != -1 || omega != -1))
die("ERROR: --transitions and --target-coverage/--expected-length cannot be used together.\n");
if ((gamma != -1 && omega == -1) || (gamma == -1 && omega != -1))
die("ERROR: --target-coverage and --expecteed-length must be used together.\n");
set_seed(-1);
if (gamma != -1)
nu = gamma/(1-gamma) * mu;
fprintf(stderr, "Reading tree model from %s...\n", argv[optind]);
source_mod = tm_new_from_file(phast_fopen(argv[optind], "r"), 1);
if (source_mod->nratecats > 1)
die("ERROR: rate variation not currently supported.\n");
if (source_mod->order > 0)
die("ERROR: only single nucleotide models are currently supported.\n");
if (!tm_is_reversible(source_mod))
phast_warning("WARNING: p-value computation assumes reversibility and your model is non-reversible.\n");
/* read alignment */
msa_f = phast_fopen(argv[optind], "r");
fprintf(stderr, "Reading alignment from %s...\n", argv[optind]);
if (msa_format == UNKNOWN_FORMAT)
msa_format = msa_format_for_content(msa_f, 1);
if (msa_format == MAF) {
msa = maf_read(msa_f, refseq_f, 1, NULL, NULL, NULL, -1, TRUE, NULL,
NO_STRIP, FALSE);
}
else
msa = msa_new_from_file_define_format(msa_f, msa_format, NULL);
if (msa_alph_has_lowercase(msa)) msa_toupper(msa);
msa_remove_N_from_alph(msa);
if (msa->ss == NULL) {
fprintf(stderr, "Extracting sufficient statistics...\n");
ss_from_msas(msa, 1, TRUE, NULL, NULL, NULL, -1, 0);
}
else if (msa->ss->tuple_idx == NULL)
die("ERROR: ordered representation of alignment required unless --suff-stats.\n");
/* prune tree, if necessary */
old_nnodes = source_mod->tree->nnodes;
tm_prune(source_mod, msa, pruned_names);
if (lst_size(pruned_names) == (old_nnodes + 1) / 2)
die("ERROR: no match for leaves of tree in alignment (leaf names must match alignment names).\n");
if (lst_size(pruned_names) > 0) {
fprintf(stderr, "WARNING: pruned away leaves of tree with no match in alignment (");
for (i = 0; i < lst_size(pruned_names); i++)
fprintf(stderr, "%s%s", ((String*)lst_get_ptr(pruned_names, i))->chars,
i < lst_size(pruned_names) - 1 ? ", " : ").\n");
}
/* this has to be done after pruning tree */
tr_name_ancestors(source_mod->tree);
/* also make sure match for reference sequence in tree */
if (refidx > 0) {
for (i = 0, found = FALSE; !found && i < source_mod->tree->nnodes; i++) {
TreeNode *n = lst_get_ptr(source_mod->tree->nodes, i);
if (!strcmp(n->name, msa->names[refidx-1]))
found = TRUE;
}
if (!found) die("ERROR: no match for reference sequence in tree.\n");
}
/* checks for indel model */
if (alpha_c > 0) {
if (ih == NULL) {
fprintf(stderr, "Reconstructing indel history by parsimony...\n");
ih = ih_reconstruct(msa, source_mod->tree);
}
else {
if (ih->ncols != msa->length)
die("ERROR: indel history doesn't seem to match alignment.\n");
if (ih->tree->nnodes != source_mod->tree->nnodes)
die("ERROR: indel history doesn't seem to match tree model.\n");
}
}
bdphmm = bd_new(source_mod, rho, mu, nu, phi, alpha_c, beta_c, tau_c,
alpha_n, beta_n, tau_n, estim_gamma, estim_omega,
estim_phi);
/* compute emissions */
phmm_compute_emissions(bdphmm->phmm, msa, FALSE);
/* add emissions for indel model, if necessary */
if (alpha_c > 0) {
fprintf(stderr, "Adjusting emissions for indels...\n");
bd_add_indel_emissions(bdphmm, ih);
}
/* postprocess for missing data (requires special handling) */
fprintf(stderr, "Adjusting emissions for missing data...\n");
bd_handle_missing_data(bdphmm, msa);
if (estim_gamma || estim_omega || estim_phi) {
fprintf(stderr, "Estimating free parameters...\n");
bd_estimate_transitions(bdphmm, msa);
}
/* set seqname and idpref, if necessary */
if (seqname == NULL || idpref == NULL) {
/* derive default from file name root */
String *tmp = str_new_charstr(msa_fname);
if (!str_equals_charstr(tmp, "-")) {
str_remove_path(tmp);
str_root(tmp, '.');
if (idpref == NULL) idpref = copy_charstr(tmp->chars);
str_root(tmp, '.'); /* apply one more time for double suffix */
if (seqname == NULL) seqname = tmp->chars;
}
else if (seqname == NULL) seqname = "refseq";
}
/* obtain predictions */
fprintf(stderr, "Running Viterbi algorithm...\n");
predictions = phmm_predict_viterbi(bdphmm->phmm, seqname, NULL, idpref, NULL);
lst_push_ptr(ignore_types, str_new_charstr("nonconserved"));
gff_filter_by_type(predictions, ignore_types, TRUE, NULL);
/* score predictions */
fprintf(stderr, "Scoring predictions...\n");
bd_score_predictions(bdphmm, predictions);
/* can free emissions now */
for (i = 0; i < bdphmm->phmm->hmm->nstates; i++)
sfree(bdphmm->phmm->emissions[i]);
sfree(bdphmm->phmm->emissions);
bdphmm->phmm->emissions = NULL;
/* convert GFF to coord frame of reference sequence and adjust
coords by idx_offset, if necessary */
if (refidx != 0 || msa->idx_offset != 0)
msa_map_gff_coords(msa, predictions, 0, refidx, msa->idx_offset);
if (refidx != 0)
gff_flatten(predictions);
/* necessary because coord conversion might create overlapping
features (can happen in deletions in reference sequence) */
/* now output predictions */
fprintf(stderr, "Writing GFF to stdout...\n");
gff_print_set(stdout, predictions);
fprintf(stderr, "Done.\n");
return 0;
}
int main(int argc, char *argv[]) {
char c;
List *l;
int i, j, strand, bed_output = 0, backgd_nmods = -1, feat_nmods = -1,
winsize = -1, verbose = 0, max_nmods, memblocksize, old_nleaves,
refidx = 1, base_by_base = FALSE, windowWig = FALSE;
TreeModel **backgd_mods = NULL, **feat_mods = NULL;
HMM *backgd_hmm = NULL, *feat_hmm = NULL;
msa_format_type inform = UNKNOWN_FORMAT;
GFF_Set *features = NULL;
MSA *msa, *msa_compl=NULL;
double **backgd_emissions, **feat_emissions, **mem, **dummy_emissions,
*winscore_pos=NULL, *winscore_neg=NULL;
int *no_alignment=NULL;
List *pruned_names;
char *msa_fname;
FILE *infile;
int opt_idx;
struct option long_opts[] = {
{"background-mods", 1, 0, 'b'},
{"background-hmm", 1, 0, 'B'},
{"feature-mods", 1, 0, 'f'},
{"feature-hmm", 1, 0, 'F'},
{"features", 1, 0, 'g'},
{"window", 1, 0, 'w'},
{"window-wig", 1, 0, 'W'},
{"base-by-base", 0, 0, 'y'},
{"msa-format", 1, 0, 'i'},
{"refidx", 1, 0, 'r'},
{"output-bed", 0, 0, 'd'},
{"verbose", 0, 0, 'v'},
{"help", 0, 0, 'h'},
{0, 0, 0, 0}
};
while ((c = getopt_long(argc, argv, "B:b:F:f:r:g:w:W:i:ydvh", long_opts, &opt_idx)) != -1) {
switch (c) {
case 'B':
backgd_hmm = hmm_new_from_file(phast_fopen(optarg, "r"));
break;
case 'b':
l = get_arg_list(optarg);
backgd_nmods = lst_size(l);
backgd_mods = smalloc(backgd_nmods * sizeof(void*));
for (i = 0; i < backgd_nmods; i++)
backgd_mods[i] = tm_new_from_file(phast_fopen(((String*)lst_get_ptr(l, i))->chars, "r"), 1);
lst_free_strings(l);
lst_free(l);
break;
case 'F':
feat_hmm = hmm_new_from_file(phast_fopen(optarg, "r"));
break;
case 'f':
l = get_arg_list(optarg);
feat_nmods = lst_size(l);
feat_mods = smalloc(feat_nmods * sizeof(void*));
for (i = 0; i < feat_nmods; i++)
feat_mods[i] = tm_new_from_file(phast_fopen(((String*)lst_get_ptr(l, i))->chars, "r"), 1);
lst_free_strings(l);
lst_free(l);
break;
case 'g':
features = gff_read_set(phast_fopen(optarg, "r"));
break;
case 'w':
winsize = get_arg_int(optarg);
if (winsize <= 0) die("ERROR: window size must be positive.\n");
break;
case 'W':
winsize = get_arg_int(optarg);
if (winsize <= 0) die("ERROR: window size must be positive.\n");
windowWig = TRUE;
break;
case 'y':
base_by_base = TRUE;
break;
case 'i':
inform = msa_str_to_format(optarg);
if (inform == UNKNOWN_FORMAT) die("Bad argument to -i.\n");
break;
case 'r':
refidx = get_arg_int_bounds(optarg, 0, INFTY);
break;
case 'd':
bed_output = 1;
break;
case 'h':
printf("%s", HELP);
exit(0);
case 'v':
verbose = 1;
break;
case '?':
die("Bad argument. Try '%s -h'.\n", argv[0]);
}
}
set_seed(-1);
if (backgd_mods == NULL || feat_mods == NULL)
die("ERROR: -b and -f required. Try '%s -h'.\n", argv[0]);
if (backgd_nmods == 1 && backgd_hmm == NULL)
backgd_hmm = hmm_create_trivial();
else if (backgd_hmm == NULL)
die("ERROR: -B required. Try '%s -h'.\n", argv[0]);
if (feat_nmods == 1 && feat_hmm == NULL)
feat_hmm = hmm_create_trivial();
else if (feat_hmm == NULL)
die("ERROR: -F required. Try '%s -h'.\n", argv[0]);
if ((winsize == -1 && features == NULL && !base_by_base) ||
(winsize != -1 && features != NULL) ||
(winsize != -1 && base_by_base) ||
(features != NULL && base_by_base))
die("ERROR: must specify exactly one of -g, -w, and -y. Try '%s -h'.\n", argv[0]);
if (backgd_hmm->nstates != backgd_nmods)
die("ERROR: number of states must equal number of tree models for background.\n");
if (feat_hmm->nstates != feat_nmods)
die("ERROR: number of states must equal number of tree models for features.\n");
if (features != NULL && lst_size(features->features) == 0)
die("ERROR: empty features file.\n");
if (base_by_base && (backgd_nmods > 1 || feat_nmods > 1))
die("ERROR: only single phylogenetic models (not HMMs) are supported with --base-by-base.\n");
if (optind != argc - 1)
die("ERROR: too few arguments. Try '%s -h'.\n", argv[0]);
if (verbose) fprintf(stderr, "Reading alignment ...\n");
msa_fname = argv[optind];
infile = phast_fopen(msa_fname, "r");
if (inform == UNKNOWN_FORMAT)
inform = msa_format_for_content(infile, 1);
if (inform == MAF)
msa = maf_read(infile, NULL, 1, NULL, NULL,
NULL, -1, TRUE, NULL, NO_STRIP, FALSE);
else
msa = msa_new_from_file_define_format(infile, inform, NULL);
if (msa_alph_has_lowercase(msa)) msa_toupper(msa);
msa_remove_N_from_alph(msa);
/* need ordered representation of alignment */
if (msa->seqs == NULL && (msa->ss == NULL || msa->ss->tuple_idx == NULL) )
die("ERROR: ordered sufficient statistics are required.\n");
pruned_names = lst_new_ptr(msa->nseqs);
for (i = 0; i < backgd_nmods; i++) {
old_nleaves = (backgd_mods[i]->tree->nnodes + 1) / 2;
tm_prune(backgd_mods[i], msa, pruned_names);
if (lst_size(pruned_names) >= old_nleaves)
die("ERROR: no match for leaves of tree in alignment (background model #%d)\n", i+1);
else if (lst_size(pruned_names) > 0) {
fprintf(stderr, "WARNING: pruned away leaves in background model (#%d) with no match in alignment (", i+1);
for (j = 0; j < lst_size(pruned_names); j++)
fprintf(stderr, "%s%s", ((String*)lst_get_ptr(pruned_names, j))->chars,
j < lst_size(pruned_names) - 1 ? ", " : ").\n");
}
lst_free_strings(pruned_names);
}
for (i = 0; i < feat_nmods; i++) {
old_nleaves = (feat_mods[i]->tree->nnodes + 1) / 2;
tm_prune(feat_mods[i], msa, pruned_names);
if (lst_size(pruned_names) >= old_nleaves)
die("ERROR: no match for leaves of tree in alignment (features model #%d)\n", i+1);
else if (lst_size(pruned_names) > 0) {
fprintf(stderr, "WARNING: pruned away leaves in features model (#%d) with no match in alignment (", i+1);
for (j = 0; j < lst_size(pruned_names); j++)
fprintf(stderr, "%s%s", ((String*)lst_get_ptr(pruned_names, j))->chars,
j < lst_size(pruned_names) - 1 ? ", " : ").\n");
}
lst_free_strings(pruned_names);
}
lst_free(pruned_names);
/* first have to subtract offset from features, if necessary */
if (msa->idx_offset != 0 && features != NULL) {
for (i = 0; i < lst_size(features->features); i++) {
GFF_Feature *f = lst_get_ptr(features->features, i);
f->start -= msa->idx_offset;
f->end -= msa->idx_offset;
}
}
/* convert to coord frame of alignment */
if (features != NULL && refidx != 0) {
if (verbose) fprintf(stderr, "Mapping coordinates ...\n");
msa_map_gff_coords(msa, features, refidx, 0, 0);
if (lst_size(features->features) == 0)
die("ERROR: no features within coordinate range of alignment.\n");
}
/* Make a reverse complemented copy of the alignment. The two
strands will be processed separately, to avoid problems with
overlapping features, etc. */
if (!base_by_base) { /* skip in base by base case */
if (verbose) fprintf(stderr, "Creating reverse complemented alignment ...\n");
msa_compl = msa_create_copy(msa, 0);
/* temporary workaround: make sure reverse complement not based on
sufficient stats */
if (msa_compl->seqs == NULL) ss_to_msa(msa_compl);
if (msa_compl->ss != NULL) {
ss_free(msa_compl->ss);
msa_compl->ss = NULL;
}
msa_reverse_compl(msa_compl);
}
/* allocate memory for computing scores */
backgd_emissions = smalloc(backgd_nmods * sizeof(void*));
for (i = 0; i < backgd_nmods; i++)
backgd_emissions[i] = smalloc(msa->length * sizeof(double));
feat_emissions = smalloc(feat_nmods * sizeof(void*));
for (i = 0; i < feat_nmods; i++)
feat_emissions[i] = smalloc(msa->length * sizeof(double));
max_nmods = max(backgd_nmods, feat_nmods);
dummy_emissions = smalloc(max_nmods * sizeof(void*));
mem = smalloc(max_nmods * sizeof(void*));
/* memory for forward algorithm -- each block must be as large as
the largest feature */
if (features != NULL) {
for (i = 0, memblocksize = -1; i < lst_size(features->features); i++) {
GFF_Feature *f = lst_get_ptr(features->features, i);
if (f->end - f->start + 1 > memblocksize)
memblocksize = f->end - f->start + 1;
}
}
else memblocksize = winsize; /* -1 if base-by-base mode */
if (memblocksize > 0)
for (i = 0; i < max_nmods; i++)
mem[i] = smalloc(memblocksize * sizeof(double));
if (winsize != -1) {
winscore_pos = smalloc(msa->length * sizeof(double));
winscore_neg = smalloc(msa->length * sizeof(double));
no_alignment = smalloc(msa->length * sizeof(int));
for (i = 0; i < msa->length; i++) {
winscore_pos[i] = winscore_neg[i] = NEGINFTY;
if (refidx == 0)
no_alignment[i] = FALSE;
else
no_alignment[i] = msa_missing_col(msa, refidx, i);
}
}
/* the rest will be repeated for each strand */
for (strand = 1; strand <= 2; strand++) {
MSA *thismsa = strand == 1 ? msa : msa_compl;
double *winscore = strand == 1 ? winscore_pos : winscore_neg;
if (base_by_base && strand == 2) break; /* don't do second pass in
base_by_base case */
if (verbose) fprintf(stderr, "Processing %c strand ...\n",
strand == 1 ? '+' : '-');
/* set up dummy categories array, so that emissions are only
computed where needed */
thismsa->categories = smalloc(thismsa->length * sizeof(int));
thismsa->ncats = 1;
if (winsize != -1) {
if (strand == 1)
for (i = 0; i < thismsa->length; i++)
thismsa->categories[i] = no_alignment[i] ? 0 : 1;
else
for (i = 0; i < thismsa->length; i++)
thismsa->categories[i] = no_alignment[thismsa->length - i - 1] ? 0 : 1;
}
else if (features != NULL) {
for (i = 0; i < thismsa->length; i++) thismsa->categories[i] = 0;
for (i = 0; i < lst_size(features->features); i++) {
GFF_Feature *f = lst_get_ptr(features->features, i);
if (f->start <= 0 || f->end <= 0) {
fprintf(stderr, "WARNING: feature out of range ('");
gff_print_feat(stderr, f);
fprintf(stderr, "')\n");
continue;
}
if (strand == 1 && f->strand != '-')
for (j = f->start - 1; j < f->end; j++)
thismsa->categories[j] = 1;
else if (strand == 2 && f->strand == '-')
for (j = thismsa->length - f->end;
j < thismsa->length - f->start + 1; j++)
thismsa->categories[j] = 1;
}
}
else { /* base-by-base scores */
for (i = 0; i < thismsa->length; i++) thismsa->categories[i] = 1;
}
if (thismsa->ss != NULL) ss_update_categories(thismsa);
/* compute emissions */
for (i = 0; i < backgd_nmods; i++) {
if (verbose)
fprintf(stderr, "Computing emissions for background model #%d ...\n", i+1);
tl_compute_log_likelihood(backgd_mods[i], thismsa,
backgd_emissions[i], NULL, 1, NULL);
}
for (i = 0; i < feat_nmods; i++) {
if (verbose)
fprintf(stderr, "Computing emissions for features model #%d ...\n", i+1);
tl_compute_log_likelihood(feat_mods[i], thismsa,
feat_emissions[i], NULL, 1, NULL);
}
/* now compute scores */
if (winsize != -1) { /* windows case */
int winstart;
if (verbose) fprintf(stderr, "Computing scores ...\n");
for (winstart = 0; winstart <= thismsa->length - winsize; winstart++) {
int centeridx = winstart + winsize/2;
if (strand == 2) centeridx = thismsa->length - centeridx - 1;
if (no_alignment[centeridx]) continue;
for (j = 0; j < feat_nmods; j++)
dummy_emissions[j] = &(feat_emissions[j][winstart]);
winscore[centeridx] = hmm_forward(feat_hmm, dummy_emissions,
winsize, mem);
if (winscore[centeridx] <= NEGINFTY) {
winscore[centeridx] = NEGINFTY;
continue;
}
for (j = 0; j < backgd_nmods; j++)
dummy_emissions[j] = &(backgd_emissions[j][winstart]);
winscore[centeridx] -= hmm_forward(backgd_hmm, dummy_emissions,
winsize, mem);
if (winscore[centeridx] < NEGINFTY) winscore[centeridx] = NEGINFTY;
}
}
else if (features != NULL) { /* features case */
if (verbose) fprintf(stderr, "Computing scores ...\n");
for (i = 0; i < lst_size(features->features); i++) {
GFF_Feature *f = lst_get_ptr(features->features, i);
int s, e;
if ((strand == 1 && f->strand == '-') ||
(strand == 2 && f->strand != '-') ||
f->start <= 0 || f->end <= 0 || f->end - f->start < 0)
continue;
/* effective coords */
if (f->strand == '-') {
s = thismsa->length - f->end + 1;
e = thismsa->length - f->start + 1;
}
else {
s = f->start;
e = f->end;
}
f->score_is_null = 0;
for (j = 0; j < feat_nmods; j++)
dummy_emissions[j] = &(feat_emissions[j][s-1]);
f->score = hmm_forward(feat_hmm, dummy_emissions, e - s + 1, mem);
if (f->score <= NEGINFTY) {
f->score = NEGINFTY;
continue;
}
for (j = 0; j < backgd_nmods; j++)
dummy_emissions[j] = &(backgd_emissions[j][s-1]);
f->score -= hmm_forward(backgd_hmm, dummy_emissions, e - s + 1, mem);
if (f->score < NEGINFTY) f->score = NEGINFTY;
}
}
}
if (verbose) fprintf(stderr, "Generating output ...\n");
if (winsize != -1 && windowWig == FALSE) { /* standard windows output */
for (i = 0, j = 0; i < msa->length; i++) {
if (no_alignment[i] == FALSE)
printf("%d\t%.3f\t%.3f\n", j + msa->idx_offset + 1, winscore_pos[i],
winscore_neg[i]);
if (ss_get_char_pos(msa, i, 0, 0) != GAP_CHAR) j++;
}
}
else if (windowWig == TRUE) { /* windows with wig output */
int last = NEGINFTY;
for (i = 0, j = 0; i < msa->length; i++) {
if (refidx == 0 || msa_get_char(msa, refidx-1, i) != GAP_CHAR) {
if (no_alignment[i] == FALSE && winscore_pos[i] > NEGINFTY) {
if (j > last + 1)
printf("fixedStep chrom=%s start=%d step=1\n",
refidx > 0 ? msa->names[refidx-1] : "alignment",
j + msa->idx_offset + 1);
printf("%.3f\n", winscore_pos[i]);
last = j;
}
j++;
}
}
}
else if (features != NULL) { /* features output */
/* return to coord frame of reference seq (also, replace offset) */
if (refidx != 0)
msa_map_gff_coords(msa, features, 0, refidx, msa->idx_offset);
else if (msa->idx_offset != 0) {
for (i = 0; i < lst_size(features->features); i++) {
GFF_Feature *f = lst_get_ptr(features->features, i);
f->start += msa->idx_offset;
f->end += msa->idx_offset;
}
}
if (bed_output)
gff_print_bed(stdout, features, FALSE);
else
gff_print_set(stdout, features);
}
else { /* base-by-base scores */
/* in this case, we can just output the difference between the emissions */
printf("fixedStep chrom=%s start=%d step=1\n",
refidx > 0 ? msa->names[refidx-1] : "alignment",
msa->idx_offset + 1);
for (i = 0, j = 0; i < msa->length; i++) {
if (refidx == 0 || msa_get_char(msa, refidx-1, i) != GAP_CHAR) {
printf("%.3f\n", feat_emissions[0][i] - backgd_emissions[0][i]);
j++;
}
}
}
if (verbose) fprintf(stderr, "\nDone.\n");
return 0;
}
int main(int argc, char *argv[]) {
TreeNode *tree = NULL;
TreeModel *backgd_mod = NULL;
int i, j,
size = DEFAULT_SIZE, meme_mode = 0, profile_mode = 0,
nrestarts = 10, npseudocounts = 5, nsamples = -1,
nmostprevalent = -1, tuple_size = -1, nbest = -1, sample_parms = 0,
nmotifs = DEFAULT_NUMBER, nseqs = -1, do_html = 0, do_bed = 0,
suppress_stdout = 0;
List *msa_name_list = NULL, *pos_examples = NULL, *init_list = NULL, *tmpl;
List *msas, *motifs;
SeqSet *seqset = NULL;
PooledMSA *pmsa = NULL;
msa_format_type msa_format = UNKNOWN_FORMAT;
Vector *backgd_mnmod = NULL;
Hashtable *hash=NULL;
String *output_prefix = str_new_charstr("phastm.");
double *has_motif = NULL;
double prior = PRIOR;
char c;
GFF_Set *bedfeats = NULL;
while ((c = getopt(argc, argv, "t:i:b:sk:md:pn:I:R:P:w:c:SB:o:HDxh")) != -1) {
switch (c) {
case 't':
tree = tr_new_from_file(phast_fopen(optarg, "r"));
break;
case 'i':
msa_format = msa_str_to_format(optarg);
if (msa_format == UNKNOWN_FORMAT)
die("ERROR: bad input format.\n");
break;
case 'b':
backgd_mod = tm_new_from_file(phast_fopen(optarg, "r"), 1);
break;
case 's':
break;
case 'k':
size = get_arg_int(optarg);
break;
case 'm':
meme_mode = 1;
break;
case 'd':
pos_examples = get_arg_list(optarg);
break;
case 'p':
profile_mode = 1;
break;
case 'n':
nrestarts = get_arg_int(optarg);
break;
case 'I':
init_list = get_arg_list(optarg);
break;
case 'P':
tmpl = str_list_as_int(get_arg_list(optarg));
if (lst_size(tmpl) != 2) die("ERROR: bad argument to -P.\n");
nmostprevalent = lst_get_int(tmpl, 0);
tuple_size = lst_get_int(tmpl, 1);
if (!(nmostprevalent > 0 && tuple_size > 0))
die("ERROR: bad argument nmostprevalent=%i tuple_size=%i\n",
nmostprevalent, tuple_size);
lst_free(tmpl);
break;
case 'R':
tmpl = str_list_as_int(get_arg_list(optarg));
if (lst_size(tmpl) != 2) die("ERROR: bad argument to -R.\n");
nsamples = lst_get_int(tmpl, 0);
tuple_size = lst_get_int(tmpl, 1);
if (!(nsamples > 0 && tuple_size > 0))
die("ERROR nsamples=%i tuple_sizse=%i\n", nsamples, tuple_size);
lst_free(tmpl);
break;
case 'c':
npseudocounts = get_arg_int(optarg);
break;
case 'w':
nbest = get_arg_int(optarg);
break;
case 'S':
sample_parms = 1;
break;
case 'B':
nmotifs = get_arg_int(optarg);
break;
case 'o':
str_free(output_prefix);
output_prefix = str_new_charstr(optarg);
str_append_char(output_prefix, '.');
break;
case 'H':
do_html = 1;
break;
case 'D':
do_bed = 1;
break;
case 'x':
suppress_stdout = 1;
break;
case 'h':
usage(argv[0]);
case '?':
die("Bad argument. Try '%s -h'.\n", argv[0]);
}
}
if (optind != argc - 1)
die("ERROR: List of alignment files required. Try '%s -h'.\n", argv[0]);
if ((nsamples > 0 && nmostprevalent > 0) ||
(nsamples > 0 && init_list != NULL) ||
(nmostprevalent > 0 && init_list != NULL))
die("ERROR: -I, -P, and -R are mutually exclusive.");
set_seed(-1);
msa_name_list = get_arg_list(argv[optind]);
if (backgd_mod != NULL && tree == NULL) tree = backgd_mod->tree;
if (tree == NULL && !meme_mode && !profile_mode)
die("ERROR: Must specify -t, -m, or -p.\n");
if ((init_list != NULL || nsamples > 0 || nmostprevalent > 0) &&
!sample_parms)
nrestarts = 1;
if (pos_examples != NULL) {
hash = hsh_new(lst_size(pos_examples));
for (i = 0; i < lst_size(pos_examples); i++)
hsh_put_int(hash, ((String*)lst_get_ptr(pos_examples, i))->chars, 1);
has_motif = smalloc(lst_size(msa_name_list) * sizeof(double));
}
/* open all MSAs */
msas = lst_new_ptr(lst_size(msa_name_list));
fprintf(stderr, "Reading alignment(s) ...\n");
for (i = 0, j = 0; i < lst_size(msa_name_list); i++) {
String *name = lst_get_ptr(msa_name_list, i);
FILE *mfile = phast_fopen(name->chars, "r");
msa_format_type temp_format;
MSA *msa;
if (msa_format == UNKNOWN_FORMAT)
temp_format = msa_format_for_content(mfile, 1);
else temp_format = msa_format;
msa = msa_new_from_file_define_format(mfile, temp_format, NULL);
phast_fclose(mfile);
if (nseqs == -1) nseqs = msa->nseqs;
if (!meme_mode &&
(msa->length - msa_num_gapped_cols(msa, STRIP_ANY_GAPS, -1, -1) < 300 ||
msa->nseqs != nseqs)) {
fprintf(stderr, "WARNING: ignoring alignment '%s' -- too few informative sites.\n", name->chars);
msa_free(msa);
continue;
}
if (msa_alph_has_lowercase(msa)) msa_toupper(msa);
msa_remove_N_from_alph(msa); /* Ns can be a problem */
lst_push_ptr(msas, msa);
if (has_motif != NULL) {
int k, hm = (hsh_get_int(hash, name->chars) == 1);
if (meme_mode) { /* here need to record at individ seq level */
has_motif = srealloc(has_motif,
(j + msa->nseqs + 1) * sizeof(double)); /* FIXME */
for (k = 0; k < msa->nseqs; k++) has_motif[j++] = hm;
}
else has_motif[j++] = hm;
}
}
if (!meme_mode) {
fprintf(stderr, "Extracting and pooling sufficient statistics ...\n");
pmsa = ss_pooled_from_msas(msas, 1, size, NULL, 0);
msa_remove_N_from_alph(pmsa->pooled_msa);
}
/* obtain individual sequences, if necessary */
if (nmostprevalent > 0 || nsamples > 0 || meme_mode) {
if (meme_mode) fprintf(stderr, "Converting to individual sequences ...\n");
else fprintf(stderr, "Obtaining reference sequences for pre-processing ...\n");
seqset = mtf_get_seqset(msas, meme_mode ? -1 : 1, 10 * size);
/* for now, assume 1st seq is reference */
msa_remove_N_from_alph(seqset->set);
}
if (nmostprevalent > 0) {
fprintf(stderr, "Obtaining %d most prevalent %d-tuples ...\n",
nmostprevalent, tuple_size);
init_list = lst_new_ptr(nmostprevalent);
mtf_get_common_ntuples(seqset, init_list, tuple_size, nmostprevalent);
}
else if (nsamples > 0) {
fprintf(stderr, "Sampling %d %d-tuples ...\n", nsamples, tuple_size);
init_list = lst_new_ptr(nsamples);
mtf_sample_ntuples(seqset, init_list, tuple_size, nsamples);
}
/* in meme_mode, backgd model can be specified as eq freqs in a .mod file */
if (meme_mode && backgd_mod != NULL && has_motif == NULL)
backgd_mnmod = backgd_mod->backgd_freqs;
/* estimate background model, if necessary */
else if (backgd_mod == NULL && (!meme_mode || has_motif == NULL)) {
fprintf(stderr, "Fitting background model%s ...\n",
has_motif == NULL ? "" : " (for use in initialization)");
/* if discriminative, be clear
backgd isn't really part of the
estimation procedure */
if (meme_mode) {
backgd_mnmod = vec_new(strlen(seqset->set->alphabet));
mtf_estim_backgd_mn(seqset, backgd_mnmod);
}
else {
backgd_mod = tm_new(tr_create_copy(tree), NULL, NULL, F81,
pmsa->pooled_msa->alphabet, 1, 0, NULL, -1);
tm_fit(backgd_mod, pmsa->pooled_msa,
tm_params_init(backgd_mod, .1, 5, 0),
-1, OPT_MED_PREC, NULL, 0, NULL);
}
}
/* select subset of init strings, if necessary */
if (nbest > 0 && init_list != NULL) {
fprintf(stderr, "Winnowing candidate start strings ...\n");
tmpl = lst_new_ptr(nbest);
mtf_winnow_starts(meme_mode ? (void*)seqset : (void*)pmsa,
init_list, nbest, tmpl, !meme_mode, size, tree,
meme_mode ? (void*)backgd_mnmod : (void*)backgd_mod,
has_motif);
lst_free(init_list);
init_list = tmpl;
}
/* Now find motifs */
motifs = mtf_find(meme_mode ? (void*)seqset : (void*)pmsa,
!meme_mode, size, nmotifs, tree,
meme_mode ? (void*)backgd_mnmod : (void*)backgd_mod,
has_motif, prior, nrestarts, init_list, sample_parms,
npseudocounts);
fprintf(stderr, "\n\n");
if (do_bed)
bedfeats = gff_new_set_init("phast_motif", "0.1b");
/* generate output */
for (i = 0; i < lst_size(motifs); i++) {
Motif *m = lst_get_ptr(motifs, i);
if (!suppress_stdout) {
if (lst_size(motifs) > 1)
printf("\n**********\nMOTIF #%d\n**********\n\n", i+1);
mtf_print(stdout, m);
}
if (do_html) {
String *fname = str_dup(output_prefix);
str_append_int(fname, i+1);
str_append_charstr(fname, ".html");
mtf_print_html(phast_fopen(fname->chars, "w+"), m);
str_free(fname);
}
if (do_bed)
mtf_add_features(m, bedfeats);
}
if (do_html) {
String *fname = str_dup(output_prefix);
str_append_charstr(fname, "index.html");
mtf_print_summary_html(phast_fopen(fname->chars, "w+"),
motifs, output_prefix);
str_free(fname);
}
if (do_bed) {
String *fname = str_dup(output_prefix);
str_append_charstr(fname, "bed");
gff_print_bed(phast_fopen(fname->chars, "w+"),
bedfeats, FALSE);
str_free(fname);
}
return 0;
}