def main(args, outs):
molecule_counter = cr_mol_counter.MoleculeCounter.open(
args.raw_molecules,
'r',
start=int(args.chunk_start),
length=int(args.chunk_len))
filtered_bcs_per_genome = cr_utils.load_barcode_csv(args.filtered_barcodes)
raw_matrices = cr_matrix.GeneBCMatrices.build_from_mol_counter(
molecule_counter)
filtered_matrices = raw_matrices.filter_barcodes(filtered_bcs_per_genome)
raw_matrices.save_h5(outs.raw_matrices_h5)
raw_matrices.save_mex(outs.raw_matrices_mex)
raw_matrices.save_barcode_summary_h5(outs.barcode_summary_h5)
filtered_matrices.save_h5(outs.filtered_matrices_h5)
filtered_matrices.save_mex(outs.filtered_matrices_mex)
genome_ids = molecule_counter.get_ref_column('genome_ids')
with cr_mol_counter.MoleculeCounter.open(outs.filtered_molecules,
'w') as ctr_out:
summary = write_filtered_molecules(molecule_counter, ctr_out,
genome_ids, filtered_bcs_per_genome)
with open(outs.summary, 'w') as f:
tk_json.dump_numpy(summary, f, pretty=True)
def get_gem_group_index_json(args, outs):
if args.gem_group_index_json:
cr_utils.copy(args.gem_group_index_json, outs.gem_group_index_json)
else:
generated_index = cr_matrix.get_gem_group_index(
args.filtered_gene_bc_matrices_h5)
if generated_index:
with open(outs.gem_group_index_json, 'w') as outfile:
tk_json.dump_numpy({"gem_group_index": generated_index},
outfile)
return outs.gem_group_index_json
def get_gem_group_index_json(args, outs):
if args.gem_group_index_json:
cr_io.copy(args.gem_group_index_json, outs.gem_group_index_json)
else:
generated_index = cr_matrix.get_gem_group_index(
args.feature_barcode_matrix)
if generated_index is not None:
with open(outs.gem_group_index_json, 'w') as outfile:
tk_json.dump_numpy({"gem_group_index": generated_index},
outfile)
else:
outs.gem_group_index_json = None
return outs.gem_group_index_json
def join(args, outs, chunk_defs, chunk_outs):
outs.corrected_bcs = [co.corrected_bcs for co in chunk_outs]
# Write barcode counts (merged by library_type)
bc_counters = cr_fastq.BarcodeCounter.merge_by(
[co.corrected_barcode_counts
for co in chunk_outs], [cd.library_type for cd in chunk_defs],
args.barcode_whitelist, args.gem_groups)
with open(outs.corrected_barcode_counts, 'w') as f:
tk_safe_json.dump_numpy(bc_counters, f)
outs.chunked_reporter = None
reporter = cr_report.merge_reporters(
[chunk_out.chunked_reporter for chunk_out in chunk_outs])
reporter.report_summary_json(outs.summary)
reporter.report_barcodes_h5(outs.barcode_summary)
def close(self):
if self.barcode_seqs:
with open(self.out_counts, 'w') as f:
tk_safe_json.dump_numpy(self.to_json(), f)
def join(args, outs, chunk_defs, chunk_outs):
outs.reads, outs.read2s, outs.tags = [], [], []
outs.gem_groups, outs.library_types, outs.library_ids, outs.read_groups = [], [], [], []
for chunk_out in chunk_outs:
outs.reads += [read for read in chunk_out.reads]
outs.read2s += [read2 for read2 in chunk_out.read2s]
outs.tags += [tags for tags in chunk_out.tags]
outs.gem_groups += [gem_group for gem_group in chunk_out.gem_groups]
outs.library_types += [lt for lt in chunk_out.library_types]
outs.library_ids += [li for li in chunk_out.library_ids]
outs.read_groups += [
read_group for read_group in chunk_out.read_groups
]
# Ensure that we have non-zero reads
if not outs.reads:
martian.exit(
"No reads found. Check the input fastqs and/or the chemistry definition"
)
# Ensure consistency of BAM comments
assert all(chunk_out.bam_comments == chunk_outs[0].bam_comments
for chunk_out in chunk_outs)
outs.bam_comments = chunk_outs[0].bam_comments
# Write barcode counts (merged by library_type)
bc_counters = BarcodeCounter.merge_by(
[co.barcode_counts
for co in chunk_outs], [cd.library_type for cd in chunk_defs],
args.barcode_whitelist, outs.gem_groups)
with open(outs.barcode_counts, 'w') as f:
tk_safe_json.dump_numpy(bc_counters, f)
# Write feature counts
feature_counts = None
for chunk_def, chunk_out in itertools.izip(chunk_defs, chunk_outs):
with open(chunk_out.feature_counts) as f:
chunk_counts = np.asarray(json.load(f), dtype=int)
if feature_counts is None:
feature_counts = chunk_counts
else:
feature_counts += chunk_counts
with open(outs.feature_counts, 'w') as f:
json.dump(tk_safe_json.json_sanitize(list(feature_counts)), f)
outs.align = cr_utils.select_alignment_params(args.align)
# Group reporters by library type
outs.chunked_reporter = None
reporter_groups = defaultdict(list)
for chunk_def, chunk_out in zip(chunk_defs, chunk_outs):
if not chunk_out.reads:
continue
chunk_lib_types = set(lt for lt in chunk_out.library_types)
assert len(chunk_lib_types) == 1
lib_type = list(chunk_lib_types)[0]
reporter_groups[lib_type].append(chunk_out.chunked_reporter)
# Merge reporters and prefix JSON keys by library type
summary = {}
for lib_type, reporters in reporter_groups.iteritems():
j = cr_report.merge_reporters(reporters).to_json()
prefix = rna_library.get_library_type_metric_prefix(lib_type)
j_prefixed = dict((prefix + k, v) for k, v in j.iteritems())
summary.update(j_prefixed)
# Use a temporary reporter to generate the metadata (w/o a prefix)
tmp_reporter = cr_report.Reporter()
tmp_reporter.store_chemistry_metadata(args.chemistry_def)
summary.update(tmp_reporter.to_json())
# Write summary JSON
with open(outs.summary, 'w') as f:
tk_safe_json.dump_numpy(summary, f, pretty=True)
def main(args, outs):
np.random.seed(0)
with cr_mol_counter.MoleculeCounter.open(args.molecules,
'r',
start=int(args.chunk_start),
length=int(
args.chunk_len)) as ctr_in:
with cr_mol_counter.MoleculeCounter.open(outs.out_molecules,
'w') as ctr_out:
metrics_in = ctr_in.get_all_metrics()
metrics_out = metrics_in.copy()
reads = ctr_in.get_column('reads')
gem_groups = ctr_in.get_column('gem_group')
if args.downsample and len(args.downsample_map) > 1:
downsample_func = np.vectorize(
lambda gem_group, read_count: np.random.binomial(
read_count, args.downsample_map[str(gem_group)][
'frac_reads_kept']))
# downsample metrics
for gg in metrics_out[cr_mol_counter.GEM_GROUPS_METRIC]:
frac_reads_kept = args.downsample_map[str(
gg)]['frac_reads_kept']
total_reads_in = metrics_in[
cr_mol_counter.GEM_GROUPS_METRIC][gg][
cr_mol_counter.GG_TOTAL_READS_METRIC]
total_reads_out = round(frac_reads_kept * total_reads_in)
metrics_out[cr_mol_counter.GEM_GROUPS_METRIC][gg][
cr_mol_counter.
GG_DOWNSAMPLED_READS_METRIC] = total_reads_out
ctr_out.set_all_metrics(metrics_out)
# downsample molecule info
subsampled_reads = downsample_func(gem_groups, reads)
for col in cr_mol_counter.MOLECULE_INFO_COLUMNS:
if col == 'reads':
data = subsampled_reads
else:
data = ctr_in.get_column(col)
ctr_out.add_many(col, data)
# pass reference info
for col in cr_mol_counter.MOLECULE_REF_COLUMNS:
ctr_out.set_ref_column(col, ctr_in.get_ref_column(col))
else:
subsampled_reads = reads
# collect summary stats
genomes = ctr_in.get_ref_column('genome_ids')
raw_conf_mapped_per_genome = {}
if len(genomes) == 1:
genome = genomes[0]
raw_conf_mapped_per_genome[genome] = subsampled_reads.sum()
else:
genome_ids = ctr_in.get_column('genome')
genome_index = cr_reference.get_genome_index(genomes)
for genome in genomes:
genome_id = cr_reference.get_genome_id(
genome, genome_index)
raw_conf_mapped_per_genome[genome] = subsampled_reads[
genome_ids == genome_id].sum()
summary = {
'raw_conf_mapped_per_genome': raw_conf_mapped_per_genome,
'mol_counter_metrics': metrics_out
}
with open(outs.summary, 'w') as f:
tk_json.dump_numpy(summary, f, pretty=True)
def main(args, outs):
reporter = vdj_report.VdjReporter()
cell_barcodes = set(vdj_utils.load_cell_barcodes_json(args.cell_barcodes))
barcode_contigs = vdj_annot.load_cell_contigs_from_json(
args.annotations, args.vdj_reference_path, group_key='barcode')
# From CDR sequence to sequence id
sequences = {}
# From clonotype (tuple of CDR ids) to clonotype id
clonotypes = {}
# From barcode to clonotype id
bc_clonotype_assignments = {}
# First pass: Just keep track of observed CDR3s
for contig_list in barcode_contigs:
# This will be a tuple of sequences like "TRA_<cdr seq>"
barcode_clonotype_tuple = contig_list.clonotype_tuple(
require_productive=not args.use_non_productive,
require_full_len=True,
require_high_conf=True)
# Give unique numerical ids to the CDR3 sequences
if barcode_clonotype_tuple:
for cdr_seq in barcode_clonotype_tuple:
sequences.setdefault(cdr_seq, len(sequences))
# From sequence id to CDR sequence
sequence_ids = {seq_id: seq for seq, seq_id in sequences.iteritems()}
# Do a second pass to potentially use non-full length contigs with a valid CDR3.
for contig_list in barcode_contigs:
if args.use_non_full_len:
barcode_clonotype_tuple = []
for c in contig_list.contigs():
(_, cl_seq) = c.clonotype_seq()
# If this contig has a CDR3 and we can infer the gene type of
# that CDR3 (either based on the contig itself or based on
# other full-length contigs that had this CDR3, then add this
# to the clonotype tuple).
if cl_seq in sequences:
# this will rescue contigs that have a chain and CDR3 assigned
# but aren't full length
barcode_clonotype_tuple.append(cl_seq)
else:
barcode_clonotype_tuple = contig_list.clonotype_tuple(
require_productive=(not args.use_non_productive),
require_full_len=True,
require_high_conf=True)
barcode_clonotype = tuple(
sorted(list(set([sequences[s] for s in barcode_clonotype_tuple]))))
if barcode_clonotype:
clonotype_id = clonotypes.setdefault(barcode_clonotype,
len(clonotypes))
bc_clonotype_assignments[contig_list.name] = clonotype_id
# From clonotype id to tuple of CDRs
clonotype_ids = {
clonotype_id: clonotype_tuple
for clonotype_tuple, clonotype_id in clonotypes.iteritems()
}
out_clonotypes = vdj_annot.report_clonotypes(reporter, 'raw',
cell_barcodes, clonotype_ids,
sequence_ids, barcode_contigs,
bc_clonotype_assignments)
with open(outs.clonotype_assignments, 'w') as out_file:
tk_safe_json.dump_numpy(tk_safe_json.json_sanitize(out_clonotypes),
out_file,
pretty=True)
# Add clonotype assignments to contig annotations
del barcode_contigs
with open(args.annotations) as f:
all_contigs = vdj_annot.load_contig_list_from_json(
f, args.vdj_reference_path)
vdj_annot.label_contigs_with_consensus(out_clonotypes, all_contigs, 'raw')
# Write augmented contig annotations
with open(outs.contig_annotations, 'w') as out_file:
vdj_annot.save_annotation_list_json(out_file, all_contigs)
with open(outs.contig_annotations_csv, 'w') as out_file:
vdj_annot.save_contig_list_csv(out_file,
all_contigs,
write_inferred=False)
with open(outs.contig_annotations_pickle, 'w') as out_file:
cPickle.dump(all_contigs, out_file, protocol=cPickle.HIGHEST_PROTOCOL)
# Write filtered contig annotations
with open(outs.filtered_contig_annotations_csv, 'w') as out_file:
filtered_contigs = filter(lambda x: x.high_confidence and x.is_cell,
all_contigs)
vdj_annot.save_contig_list_csv(out_file,
filtered_contigs,
write_inferred=False)
# Set a default value for paired clonotype diversity so that it will be
# present in the metric summary csv even when there are no paired cells
# or in denovo mode
paired_diversity_metric = reporter._get_metric_attr(
'vdj_paired_clonotype_diversity', MULTI_REFS_PREFIX, 'raw')
if not paired_diversity_metric.d:
paired_diversity_metric.add(None, 0)
reporter.report_summary_json(outs.summary)
