def add_vcf_dataset(alignment_group, vcf_dataset_type, vcf_output_filename):
"""
Sort the vcf file, and create a vcf dataset, Add it to the alignment group.
"""
sort_vcf(vcf_output_filename)
# If a Dataset already exists, delete it, might have been a bad run.
existing_set = Dataset.objects.filter(
type=vcf_dataset_type,
label=vcf_dataset_type,
filesystem_location=clean_filesystem_location(
vcf_output_filename)
)
if len(existing_set) > 0:
existing_set[0].delete()
vcf_dataset = Dataset.objects.create(
type=vcf_dataset_type,
label=vcf_dataset_type,
filesystem_location=clean_filesystem_location(
vcf_output_filename),
)
alignment_group.dataset_set.add(vcf_dataset)
return vcf_dataset
def add_vcf_dataset(alignment_group, vcf_dataset_type, vcf_output_filename):
"""Sort vcf file, creates vcf dataset, and adds it to the alignment group.
"""
if not os.path.exists(vcf_output_filename):
return None
sort_vcf(vcf_output_filename)
# If a Dataset already exists, delete it, might have been a bad run.
existing_set = Dataset.objects.filter(
type=vcf_dataset_type,
label=vcf_dataset_type,
filesystem_location=clean_filesystem_location(vcf_output_filename))
if len(existing_set) > 0:
existing_set[0].delete()
vcf_dataset = Dataset.objects.create(
type=vcf_dataset_type,
label=vcf_dataset_type,
filesystem_location=clean_filesystem_location(vcf_output_filename),
)
alignment_group.dataset_set.add(vcf_dataset)
return vcf_dataset
def test_dataset_compression_piping(self):
"""
Make sure data set compression behaves correctly.
"""
dataset = Dataset.objects.create(label='test_dataset',
type=Dataset.TYPE.FASTQ1)
GZIPPED_FASTQ_FILEPATH = os.path.join(settings.PWD, 'test_data',
'compressed_fastq',
'sample0.simLibrary.1.fq.gz')
dataset.filesystem_location = clean_filesystem_location(
GZIPPED_FASTQ_FILEPATH)
assert dataset.is_compressed()
process = subprocess.Popen(
('head ' + dataset.wrap_if_compressed() + ' | wc -l'),
shell=True,
executable=settings.BASH_PATH,
stdout=subprocess.PIPE,
stderr=subprocess.PIPE)
wc_output, errmsg = process.communicate()
rc = process.returncode
assert rc == 0, (
"Compression process returned non-zero exit status: %s" % (errmsg))
assert int(wc_output) == 10, ("Compression failed: %s" % (errmsg))
def test_clean_filesystem_location(self):
FAKE_ABS_ROOT = '/root/of/all/evil'
EXPECTED_CLEAN_URL = 'projects/blah'
dirty_full_url = os.path.join(FAKE_ABS_ROOT, settings.MEDIA_ROOT,
EXPECTED_CLEAN_URL)
clean_location = clean_filesystem_location(dirty_full_url)
self.assertEqual(EXPECTED_CLEAN_URL, clean_location)
def test_clean_filesystem_location(self):
FAKE_ABS_ROOT = '/root/of/all/evil'
EXPECTED_CLEAN_URL = 'projects/blah'
dirty_full_url = os.path.join(FAKE_ABS_ROOT, settings.MEDIA_ROOT,
EXPECTED_CLEAN_URL)
clean_location = clean_filesystem_location(dirty_full_url)
self.assertEqual(EXPECTED_CLEAN_URL, clean_location)
def test_multiple_chromosome_dataset_import(self):
user = User.objects.create_user(
TEST_USERNAME, password=TEST_PASSWORD, email=TEST_EMAIL)
project = Project.objects.create(
title=TEST_PROJECT_NAME, owner=user.get_profile())
test_yeast_genome = ReferenceGenome.objects.create(
project=project,
label='superbrewer2000')
test_dataset_path = os.path.join(settings.PWD, 'test_data/yeast_chrom_jkl.fasta')
dataset_path = copy_dataset_to_entity_data_dir(test_yeast_genome, test_dataset_path)
test_chroms_dataset = Dataset.objects.create(
label='jkl_chroms',
type=Dataset.TYPE.REFERENCE_GENOME_FASTA,
filesystem_location=clean_filesystem_location(dataset_path))
test_yeast_genome.dataset_set.add(test_chroms_dataset)
# Assert correct number of chromosomes
assert(test_yeast_genome.num_chromosomes == 3)
# Assert correct number of bases
assert(test_yeast_genome.num_bases == sum([chrom.num_bases for chrom in
Chromosome.objects.filter(reference_genome=test_yeast_genome)]))
# Assert correct chromosome labels
expected_chrom_names = [
'gi|448092123|ref|NC_020215.1|',
'gi|448096713|ref|NC_020216.1|',
'gi|448100869|ref|NC_020217.1|']
assert([chrom.label for chrom in Chromosome.objects.filter(reference_genome=test_yeast_genome)] == expected_chrom_names)
def test_dataset_compression_piping(self):
"""
Make sure data set compression behaves correctly.
"""
dataset = Dataset.objects.create(
label='test_dataset',
type=Dataset.TYPE.FASTQ1)
GZIPPED_FASTQ_FILEPATH = os.path.join(settings.PWD, 'test_data',
'compressed_fastq', 'sample0.simLibrary.1.fq.gz')
dataset.filesystem_location = clean_filesystem_location(
GZIPPED_FASTQ_FILEPATH)
assert dataset.is_compressed()
process = subprocess.Popen(
('head '+dataset.wrap_if_compressed()+' | wc -l'),
shell=True, executable=settings.BASH_PATH, stdout=subprocess.PIPE,
stderr=subprocess.PIPE)
wc_output, errmsg = process.communicate()
rc = process.returncode
assert rc == 0, (
"Compression process returned non-zero exit status: %s" % (
errmsg))
assert int(wc_output) == 10, (
"Compression failed: %s" % (errmsg))
def compute_callable_loci(reference_genome, sample_alignment,
bam_file_location, stderr=None):
# Set output fn to None in case try fails.
callable_loci_bed_fn = None
try:
ref_genome_fasta_location = get_dataset_with_type(
reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
output = _get_callable_loci_output_filename(bam_file_location)
get_callable_loci(bam_file_location, output)
# Add callable loci bed as dataset
callable_loci_bed = Dataset.objects.create(
label=Dataset.TYPE.BED_CALLABLE_LOCI,
type=Dataset.TYPE.BED_CALLABLE_LOCI,
filesystem_location=clean_filesystem_location(output))
sample_alignment.dataset_set.add(callable_loci_bed)
sample_alignment.save()
callable_loci_bed_fn = callable_loci_bed.get_absolute_location()
output = subprocess.check_output(
['cat', callable_loci_bed_fn])
with open(callable_loci_bed_fn, 'w') as callable_loci_bed_fh:
for i, line in enumerate(output.split('\n')):
try:
fields = line.split()
if len(fields) == 0:
continue
chrom, start, end, feature = fields
feature = titlecase_spaces(feature)
# Bed feature can't have spaces =(
feature = feature.replace(' ', '_')
print >> callable_loci_bed_fh, '\t'.join(
[chrom, start, end, feature])
except Exception as e:
print >> stderr, (
'WARNING: Callable Loci line' +
'%d: (%s) couldn\'t be parsed: %s') % (
i, line, str(e))
# add it as a jbrowse track
add_bed_file_track(reference_genome, sample_alignment, callable_loci_bed)
except Exception as e:
print >> stderr, 'WARNING: Callable Loci failed.'
print >> stderr, str(e)
return callable_loci_bed_fn
def get_split_reads(sample_alignment):
"""Isolate split reads from a sample alignment.
This uses a python script supplied with Lumpy that is run as a
separate process.
NOTE THAT THIS SCRIPT ONLY WORKS WITH BWA MEM.
"""
bwa_split_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_SPLIT)
if bwa_split_dataset is not None:
if (bwa_split_dataset.status == Dataset.STATUS.READY
and os.path.exists(bwa_split_dataset.get_absolute_location())):
return bwa_split_dataset
else:
bwa_split_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_SPLIT,
type=Dataset.TYPE.BWA_SPLIT,
status=Dataset.STATUS.NOT_STARTED)
sample_alignment.dataset_set.add(bwa_split_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_split_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_split_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(
bam_filename), "BAM file '%s' is missing." % (bam_filename)
bam_split_filename = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_split_reads.bam')
try:
bwa_split_dataset.status = Dataset.STATUS.COMPUTING
bwa_split_dataset.save(update_fields=['status'])
extract_split_reads(bam_filename, bam_split_filename)
except subprocess.CalledProcessError:
# if there are no split reads, then fail.
bwa_split_dataset.filesystem_location = ''
bwa_split_dataset.status = Dataset.STATUS.FAILED
finally:
bwa_split_dataset.status = Dataset.STATUS.READY
bwa_split_dataset.filesystem_location = clean_filesystem_location(
bam_split_filename)
bwa_split_dataset.save()
return bwa_split_dataset
def get_split_reads(sample_alignment):
"""Isolate split reads from a sample alignment.
This uses a python script supplied with Lumpy that is run as a
separate process.
NOTE THAT THIS SCRIPT ONLY WORKS WITH BWA MEM.
"""
bwa_split_dataset = get_dataset_with_type(
sample_alignment, Dataset.TYPE.BWA_SPLIT)
if bwa_split_dataset is not None:
if (bwa_split_dataset.status == Dataset.STATUS.READY and
os.path.exists(bwa_split_dataset.get_absolute_location())):
return bwa_split_dataset
else:
bwa_split_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_SPLIT,
type=Dataset.TYPE.BWA_SPLIT,
status=Dataset.STATUS.NOT_STARTED)
sample_alignment.dataset_set.add(bwa_split_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_split_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_split_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment, Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(bam_filename), "BAM file '%s' is missing." % (
bam_filename)
bam_split_filename = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_split_reads.bam')
try:
bwa_split_dataset.status = Dataset.STATUS.COMPUTING
bwa_split_dataset.save(update_fields=['status'])
extract_split_reads(bam_filename, bam_split_filename)
except subprocess.CalledProcessError:
# if there are no split reads, then fail.
bwa_split_dataset.filesystem_location = ''
bwa_split_dataset.status = Dataset.STATUS.FAILED
finally:
bwa_split_dataset.status = Dataset.STATUS.READY
bwa_split_dataset.filesystem_location = clean_filesystem_location(
bam_split_filename)
bwa_split_dataset.save()
return bwa_split_dataset
def get_discordant_read_pairs(sample_alignment):
"""Isolate discordant pairs of reads from a sample alignment.
"""
# First, check if completed dataset already exists.
bwa_disc_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_DISCORDANT)
if bwa_disc_dataset is not None:
if (bwa_disc_dataset.status == Dataset.STATUS.READY
and os.path.exists(bwa_disc_dataset.get_absolute_location())):
return bwa_disc_dataset
else:
bwa_disc_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_DISCORDANT,
type=Dataset.TYPE.BWA_DISCORDANT)
sample_alignment.dataset_set.add(bwa_disc_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_disc_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_disc_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(
bam_filename), "BAM file '%s' is missing." % (bam_filename)
# NOTE: This assumes the index just adds at .bai, w/ same path otherwise
# - will this always be true?
if not os.path.exists(bam_filename + '.bai'):
index_bam_file(bam_filename)
bam_discordant_filename = os.path.join(
sample_alignment.get_model_data_dir(), 'bwa_discordant_pairs.bam')
try:
bwa_disc_dataset.status = Dataset.STATUS.COMPUTING
bwa_disc_dataset.save(update_fields=['status'])
extract_discordant_read_pairs(bam_filename, bam_discordant_filename)
except subprocess.CalledProcessError:
bwa_disc_dataset.filesystem_location = ''
bwa_disc_dataset.status = Dataset.STATUS.FAILED
finally:
bwa_disc_dataset.status = Dataset.STATUS.READY
bwa_disc_dataset.filesystem_location = clean_filesystem_location(
bam_discordant_filename)
bwa_disc_dataset.save()
return bwa_disc_dataset
def get_discordant_read_pairs(sample_alignment):
"""Isolate discordant pairs of reads from a sample alignment.
"""
# First, check if completed dataset already exists.
bwa_disc_dataset = get_dataset_with_type(
sample_alignment, Dataset.TYPE.BWA_DISCORDANT)
if bwa_disc_dataset is not None:
if (bwa_disc_dataset.status == Dataset.STATUS.READY and
os.path.exists(bwa_disc_dataset.get_absolute_location())):
return bwa_disc_dataset
else:
bwa_disc_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_DISCORDANT,
type=Dataset.TYPE.BWA_DISCORDANT)
sample_alignment.dataset_set.add(bwa_disc_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_disc_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_disc_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment, Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(bam_filename), "BAM file '%s' is missing." % (
bam_filename)
# NOTE: This assumes the index just adds at .bai, w/ same path otherwise
# - will this always be true?
if not os.path.exists(bam_filename+'.bai'):
index_bam_file(bam_filename)
bam_discordant_filename = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_discordant_pairs.bam')
try:
bwa_disc_dataset.status = Dataset.STATUS.COMPUTING
bwa_disc_dataset.save(update_fields=['status'])
extract_discordant_read_pairs(bam_filename, bam_discordant_filename)
except subprocess.CalledProcessError:
bwa_disc_dataset.filesystem_location = ''
bwa_disc_dataset.status = Dataset.STATUS.FAILED
finally:
bwa_disc_dataset.status = Dataset.STATUS.READY
bwa_disc_dataset.filesystem_location = clean_filesystem_location(
bam_discordant_filename)
bwa_disc_dataset.save()
return bwa_disc_dataset
def add_dataset_to_entity(entity, dataset_label, dataset_type,
filesystem_location=None):
"""Helper function for adding a Dataset to a model.
"""
dataset = Dataset.objects.create(
label=dataset_label, type=dataset_type)
if filesystem_location is not None:
dataset.filesystem_location = clean_filesystem_location(
filesystem_location)
dataset.save()
entity.dataset_set.add(dataset)
entity.save()
return dataset
def add_dataset_to_entity(entity, dataset_label, dataset_type,
filesystem_location=None):
"""Helper function for adding a Dataset to a model.
"""
dataset = Dataset.objects.create(
label=dataset_label, type=dataset_type)
if filesystem_location is not None:
dataset.filesystem_location = clean_filesystem_location(
filesystem_location)
dataset.save()
entity.dataset_set.add(dataset)
entity.save()
return dataset
def derivation_fn(sample_alignment, unmapped_reads_dataset):
# Get the original bam file.
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
# Allocate a filename for the unmapped reads.
unmapped_reads_bam_file = (os.path.splitext(bam_filename)[0] +
'.unmapped.bam')
unmapped_reads_dataset.filesystem_location = clean_filesystem_location(
unmapped_reads_bam_file)
unmapped_reads_dataset.save(update_fields=['filesystem_location'])
cmd = '{samtools} view -h -b -f 0x4 {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY, bam_filename=bam_filename)
with open(unmapped_reads_bam_file, 'w') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
def derivation_fn(sample_alignment, unmapped_reads_dataset):
# Get the original bam file.
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
# Allocate a filename for the unmapped reads.
unmapped_reads_bam_file = (os.path.splitext(bam_filename)[0] +
'.unmapped.bam')
unmapped_reads_dataset.filesystem_location = clean_filesystem_location(
unmapped_reads_bam_file)
unmapped_reads_dataset.save(update_fields=['filesystem_location'])
cmd = '{samtools} view -h -b -f 0x4 {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=bam_filename)
with open(unmapped_reads_bam_file, 'w') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
def main():
# Create a User and Project.
user = get_or_create_user()
test_project = Project.objects.create(title=EXAMPLE_PROJECT_NAME,
owner=user.get_profile())
ref_genome = import_reference_genome_from_local_file(test_project,
'mg1655',
MG1655_REF_GENOME,
'genbank',
move=False)
# Create alignment group and and relate the vcf Dataset to it.
alignment_group = AlignmentGroup.objects.create(
label='Fix Recoli Alignment',
reference_genome=ref_genome,
aligner=AlignmentGroup.ALIGNER.BWA)
vcf_output_path = get_snpeff_vcf_output_path(alignment_group,
Dataset.TYPE.BWA_ALIGN)
shutil.copy(LARGE_VCF, vcf_output_path)
dataset = Dataset.objects.create(
type=Dataset.TYPE.VCF_FREEBAYES_SNPEFF,
label=Dataset.TYPE.VCF_FREEBAYES_SNPEFF,
filesystem_location=clean_filesystem_location(vcf_output_path),
)
alignment_group.dataset_set.add(dataset)
# Import ExperimentSampleo objects, setting specific uid to match
# the vcf file.
with open(EXPERIMENT_SAMPLE_MODEL_DATA_PICKLE) as sample_data_fh:
es_data = pickle.load(sample_data_fh)
for es in es_data:
es_obj = ExperimentSample.objects.create(uid=es.uid,
project=test_project,
label=es.label)
es_obj.data.update({
'group': es.group,
'well': es.well,
'num_reads': es.num_reads
})
es_obj.save()
parse_alignment_group_vcf(alignment_group,
Dataset.TYPE.VCF_FREEBAYES_SNPEFF)
def main():
# Create a User and Project.
user = get_or_create_user()
test_project = Project.objects.create(
title=EXAMPLE_PROJECT_NAME, owner=user.get_profile())
ref_genome = import_reference_genome_from_local_file(test_project,
'mg1655', MG1655_REF_GENOME, 'genbank', move=False)
# Create alignment group and and relate the vcf Dataset to it.
alignment_group = AlignmentGroup.objects.create(
label='Fix Recoli Alignment',
reference_genome=ref_genome,
aligner=AlignmentGroup.ALIGNER.BWA)
vcf_output_path = get_snpeff_vcf_output_path(alignment_group,
Dataset.TYPE.BWA_ALIGN)
shutil.copy(LARGE_VCF, vcf_output_path)
dataset = Dataset.objects.create(
type=Dataset.TYPE.VCF_FREEBAYES_SNPEFF,
label=Dataset.TYPE.VCF_FREEBAYES_SNPEFF,
filesystem_location=clean_filesystem_location(vcf_output_path),
)
alignment_group.dataset_set.add(dataset)
# Import ExperimentSampleo objects, setting specific uid to match
# the vcf file.
with open(EXPERIMENT_SAMPLE_MODEL_DATA_PICKLE) as sample_data_fh:
es_data = pickle.load(sample_data_fh)
for es in es_data:
es_obj = ExperimentSample.objects.create(
uid=es.uid,
project=test_project,
label=es.label
)
es_obj.data.update(
{'group':es.group,
'well':es.well,
'num_reads':es.num_reads})
es_obj.save()
parse_alignment_group_vcf(alignment_group,
Dataset.TYPE.VCF_FREEBAYES_SNPEFF)
def compute_callable_loci(reference_genome,
sample_alignment,
bam_file_location,
stderr=None):
# Set output fn to None in case try fails.
callable_loci_bed_fn = None
try:
ref_genome_fasta_location = get_dataset_with_type(
reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
callable_loci_bed_fn = (
_get_callable_loci_output_filename(bam_file_location))
get_callable_loci(bam_file_location, callable_loci_bed_fn)
# Add callable loci bed as dataset
callable_loci_bed_dataset = Dataset.objects.create(
label=Dataset.TYPE.BED_CALLABLE_LOCI,
type=Dataset.TYPE.BED_CALLABLE_LOCI,
filesystem_location=clean_filesystem_location(
callable_loci_bed_fn))
sample_alignment.dataset_set.add(callable_loci_bed_dataset)
sample_alignment.save()
clean_bed_fn = clean_bed_features(callable_loci_bed_dataset,
stderr=stderr)
# add it as a jbrowse track
add_bed_file_track(reference_genome, sample_alignment,
callable_loci_bed_dataset)
except Exception as e:
print >> stderr, 'WARNING: Callable Loci failed.'
print >> stderr, str(e)
clean_bed_fn = ''
finally:
return clean_bed_fn
def compute_callable_loci(reference_genome, sample_alignment,
bam_file_location, stderr=None):
# Set output fn to None in case try fails.
callable_loci_bed_fn = None
try:
ref_genome_fasta_location = get_dataset_with_type(
reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
callable_loci_bed_fn = (
_get_callable_loci_output_filename(bam_file_location))
get_callable_loci(bam_file_location, callable_loci_bed_fn)
# Add callable loci bed as dataset
callable_loci_bed_dataset = Dataset.objects.create(
label=Dataset.TYPE.BED_CALLABLE_LOCI,
type=Dataset.TYPE.BED_CALLABLE_LOCI,
filesystem_location=clean_filesystem_location(callable_loci_bed_fn))
sample_alignment.dataset_set.add(callable_loci_bed_dataset)
sample_alignment.save()
clean_bed_fn = clean_bed_features(callable_loci_bed_dataset, stderr=stderr)
# add it as a jbrowse track
add_bed_file_track(
reference_genome,
sample_alignment,
callable_loci_bed_dataset)
except Exception as e:
print >> stderr, 'WARNING: Callable Loci failed.'
print >> stderr, str(e)
clean_bed_fn = ''
finally:
return clean_bed_fn
def test_multiple_chromosome_dataset_import(self):
user = User.objects.create_user(TEST_USERNAME,
password=TEST_PASSWORD,
email=TEST_EMAIL)
project = Project.objects.create(title=TEST_PROJECT_NAME,
owner=user.get_profile())
test_yeast_genome = ReferenceGenome.objects.create(
project=project, label='superbrewer2000')
test_dataset_path = os.path.join(settings.PWD,
'test_data/yeast_chrom_jkl.fasta')
dataset_path = copy_dataset_to_entity_data_dir(test_yeast_genome,
test_dataset_path)
test_chroms_dataset = Dataset.objects.create(
label='jkl_chroms',
type=Dataset.TYPE.REFERENCE_GENOME_FASTA,
filesystem_location=clean_filesystem_location(dataset_path))
test_yeast_genome.dataset_set.add(test_chroms_dataset)
# Assert correct number of chromosomes
assert (test_yeast_genome.num_chromosomes == 3)
# Assert correct number of bases
assert (test_yeast_genome.num_bases == sum([
chrom.num_bases for chrom in Chromosome.objects.filter(
reference_genome=test_yeast_genome)
]))
# Assert correct chromosome labels
expected_chrom_names = [
'gi|448092123|ref', 'gi|448096713|ref', 'gi|448100869|ref'
]
assert (set([
chrom.seqrecord_id for chrom in Chromosome.objects.filter(
reference_genome=test_yeast_genome)
]) == set(expected_chrom_names))
def sanitize_sequence_dataset(dataset):
dataset_type_to_parse_format = {
Dataset.TYPE.REFERENCE_GENOME_FASTA: 'fasta',
Dataset.TYPE.REFERENCE_GENOME_GENBANK: 'genbank'
}
if dataset.type not in dataset_type_to_parse_format:
return
dirty_file_path = dataset.get_absolute_location()
parse_format = dataset_type_to_parse_format[dataset.type]
needs_santizing = False
with open(dirty_file_path, 'r') as dirty_fh:
for seq_record in SeqIO.parse(dirty_fh, parse_format):
if len(seq_record.id) > 16:
needs_santizing = True
break
if not needs_santizing:
return
prefix, ext = os.path.splitext(dirty_file_path)
clean_file_path = prefix + '.clean' + ext
seq_record_list = []
with open(dirty_file_path, 'r') as dirty_fh:
for seq_record in SeqIO.parse(dirty_fh, parse_format):
seq_record.id = seq_record.id[:16]
seq_record.name = seq_record.id
seq_record_list.append(seq_record)
with open(clean_file_path, 'w') as clean_fh:
SeqIO.write(seq_record_list, clean_fh, parse_format)
dataset.filesystem_location = clean_filesystem_location(clean_file_path)
dataset.save()
def sanitize_sequence_dataset(dataset):
dataset_type_to_parse_format = {
Dataset.TYPE.REFERENCE_GENOME_FASTA: 'fasta',
Dataset.TYPE.REFERENCE_GENOME_GENBANK: 'genbank'
}
if dataset.type not in dataset_type_to_parse_format:
return
dirty_file_path = dataset.get_absolute_location()
parse_format = dataset_type_to_parse_format[dataset.type]
needs_santizing = False
with open(dirty_file_path, 'r') as dirty_fh:
for seq_record in SeqIO.parse(dirty_fh, parse_format):
if len(seq_record.id) > 16:
needs_santizing = True
break
if not needs_santizing:
return
prefix, ext = os.path.splitext(dirty_file_path)
clean_file_path = prefix + '.clean' + ext
seq_record_list = []
with open(dirty_file_path, 'r') as dirty_fh:
for seq_record in SeqIO.parse(dirty_fh, parse_format):
seq_record.id = seq_record.id[:16]
seq_record.name = seq_record.id
seq_record_list.append(seq_record)
with open(clean_file_path, 'w') as clean_fh:
SeqIO.write(seq_record_list, clean_fh, parse_format)
dataset.filesystem_location = clean_filesystem_location(clean_file_path)
dataset.save()
def test_run_lumpy(self):
TEST_SAMPLE_UID = '8c57e7b9'
# Create a ref genome.
self.reference_genome = import_reference_genome_from_local_file(
self.project, 'ref_genome', TEST_FASTA, 'fasta')
# Create a sample.
self.experiment_sample = ExperimentSample.objects.create(
uid=TEST_SAMPLE_UID, project=self.project, label='sample1')
# Create a new alignment group.
alignment_group = AlignmentGroup.objects.create(
label='test alignment', reference_genome=self.reference_genome)
self.alignment_group = alignment_group
# Create the expected models.
sample_alignment = ExperimentSampleToAlignment.objects.create(
alignment_group=alignment_group,
experiment_sample=self.experiment_sample)
bwa_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_ALIGN,
type=Dataset.TYPE.BWA_ALIGN,
status=Dataset.STATUS.READY)
bwa_dataset.filesystem_location = clean_filesystem_location(
TEST_DISC_SPLIT_BAM)
bwa_dataset.save()
sample_alignment.dataset_set.add(bwa_dataset)
sample_alignment.save()
self.bwa_dataset = bwa_dataset
self.sample_alignment = sample_alignment
fasta_ref = get_dataset_with_type(
self.reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
sample_alignments = [self.sample_alignment]
vcf_output_dir = self.alignment_group.get_model_data_dir()
vcf_output_filename = os.path.join(vcf_output_dir, 'lumpy.vcf')
alignment_type = 'BWA_ALIGN'
# NOTE: Running these functions but not checking results.
get_discordant_read_pairs(self.sample_alignment)
get_split_reads(self.sample_alignment)
run_lumpy(fasta_ref, sample_alignments, vcf_output_dir,
vcf_output_filename, alignment_type)
dataset = Dataset.objects.create(
type=Dataset.TYPE.VCF_LUMPY,
label=Dataset.TYPE.VCF_LUMPY,
filesystem_location=vcf_output_filename,
)
self.alignment_group.dataset_set.add(dataset)
# Parse the resulting vcf, grab variant objects
parse_alignment_group_vcf(self.alignment_group, Dataset.TYPE.VCF_LUMPY)
# Grab the resulting variants.
variants = Variant.objects.filter(reference_genome=self.reference_genome)
# There should be a Variant object for each sv event.
self.assertEqual(2, len(variants))
# One event should be located very close to 25k
va_positions = [v.position for v in variants]
va_offset = [25000 - va_pos for va_pos in va_positions]
self.assertTrue(any([v < 50 for v in va_offset]))
def derivation_fn(sample_alignment, new_dataset):
"""Creates a bam file with reads relevant for de novo assembly.
"""
# Get the original bam file.
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
orig_bam_filename = bam_dataset.get_absolute_location()
# Allocate a filename for the new dataset.
de_novo_bam_filelocation = (os.path.splitext(orig_bam_filename)[0] +
'.de_novo.bam')
new_dataset.filesystem_location = clean_filesystem_location(
de_novo_bam_filelocation)
new_dataset.save(update_fields=['filesystem_location'])
### Strategy
# 0. Create intermediate sam
# 1. Grab unmapped reads.
# a. If the corresponding pair for any read was mapped (and thus not
# in the unmapped dataset), grab that from the original bam file.
# 2. Grab split reads.
# 2b. Add reads that are in intervals of interest.
# 3. Sort by name so that paired reads are next to each other
# 4. Filter out duplicate reads. Requires sort in step 3.
# 5. Convert to bam.
# 6. Delete intermediate files.
# 0. Create intermediate files.
intermediate_sam = (os.path.splitext(de_novo_bam_filelocation)[0] +
'.int.sam')
paired_intermediate_sam = (os.path.splitext(intermediate_sam)[0] +
'.paired.sam')
sorted_intermediate_sam = (
os.path.splitext(paired_intermediate_sam)[0] + '.sorted.sam')
deduped_sorted_intermediate_sam = (
os.path.splitext(sorted_intermediate_sam)[0] + '.deduped.sam')
intermediate_files = [
intermediate_sam, paired_intermediate_sam, sorted_intermediate_sam,
deduped_sorted_intermediate_sam
]
# 1. Get unmapped reads.
unmapped_reads_dataset = get_unmapped_reads(sample_alignment)
unmapped_reads_bam_file = unmapped_reads_dataset.get_absolute_location(
)
cmd = '{samtools} view -h {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=unmapped_reads_bam_file)
with open(intermediate_sam, 'w') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
# 2. Grab split reads.
split_reads_dataset = get_split_reads(sample_alignment)
split_rads_bam_file = split_reads_dataset.get_absolute_location()
cmd = '{samtools} view {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=split_rads_bam_file)
with open(intermediate_sam, 'a') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
# 2b. Add reads that are in any of intervals that we want to include.
if force_include_reads_in_intervals:
with open(intermediate_sam, 'a') as output_fh:
get_reads_in_interval_list(orig_bam_filename,
force_include_reads_in_intervals,
output_fh)
# 2c. For each of the reads that we've included, grab their
# corresponding pairs.
add_paired_mates(intermediate_sam, orig_bam_filename,
paired_intermediate_sam)
# 3. Sort by name so that paired reads are next to each other
cmd = ('(grep ^"@" {sam_file}; grep -v ^"@" {sam_file} | '
'sort -k1,1 -k2,2n) > {sorted_sam_file}').format(
sam_file=paired_intermediate_sam,
sorted_sam_file=sorted_intermediate_sam)
subprocess.call(cmd, shell=True)
# 4. Filter out duplicate reads. Requires sort in step 3.
cmd = 'uniq {sorted_sam_file} > {deduped_sam_file}'.format(
sorted_sam_file=sorted_intermediate_sam,
deduped_sam_file=deduped_sorted_intermediate_sam)
subprocess.call(cmd, shell=True)
# 5. Convert to bam.
cmd = '{samtools} view -Sb {sam_file} > {final_bam_file}'.format(
samtools=settings.SAMTOOLS_BINARY,
sam_file=intermediate_sam,
final_bam_file=de_novo_bam_filelocation)
subprocess.call(cmd, shell=True)
# 6. Delete intermediate files.
for f in intermediate_files:
os.remove(f)
def test_run_lumpy(self):
TEST_SAMPLE_UID = '8c57e7b9'
# Create a ref genome.
self.reference_genome = import_reference_genome_from_local_file(
self.project, 'ref_genome', TEST_FASTA, 'fasta')
# Create a sample.
self.experiment_sample = ExperimentSample.objects.create(
uid=TEST_SAMPLE_UID, project=self.project, label='sample1')
# Create a new alignment group.
alignment_group = AlignmentGroup.objects.create(
label='test alignment', reference_genome=self.reference_genome)
self.alignment_group = alignment_group
# Create the expected models.
sample_alignment = ExperimentSampleToAlignment.objects.create(
alignment_group=alignment_group,
experiment_sample=self.experiment_sample)
bwa_dataset = Dataset.objects.create(label=Dataset.TYPE.BWA_ALIGN,
type=Dataset.TYPE.BWA_ALIGN,
status=Dataset.STATUS.READY)
bwa_dataset.filesystem_location = clean_filesystem_location(
TEST_DISC_SPLIT_BAM)
bwa_dataset.save()
sample_alignment.dataset_set.add(bwa_dataset)
sample_alignment.save()
self.bwa_dataset = bwa_dataset
self.sample_alignment = sample_alignment
fasta_ref = get_dataset_with_type(
self.reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
sample_alignments = [self.sample_alignment]
vcf_output_dir = self.alignment_group.get_model_data_dir()
vcf_output_filename = os.path.join(vcf_output_dir, 'lumpy.vcf')
alignment_type = 'BWA_ALIGN'
# NOTE: Running these functions but not checking results.
get_discordant_read_pairs(self.sample_alignment)
get_split_reads(self.sample_alignment)
run_lumpy(fasta_ref, sample_alignments, vcf_output_dir,
vcf_output_filename, alignment_type)
dataset = Dataset.objects.create(
type=Dataset.TYPE.VCF_LUMPY,
label=Dataset.TYPE.VCF_LUMPY,
filesystem_location=vcf_output_filename,
)
self.alignment_group.dataset_set.add(dataset)
# Parse the resulting vcf, grab variant objects
parse_alignment_group_vcf(self.alignment_group, Dataset.TYPE.VCF_LUMPY)
# Grab the resulting variants.
variants = Variant.objects.filter(
reference_genome=self.reference_genome)
# There should be a Variant object for each sv event.
self.assertEqual(2, len(variants))
# One event should be located very close to 25k
va_positions = [v.position for v in variants]
va_offset = [25000 - va_pos for va_pos in va_positions]
self.assertTrue(any([v < 50 for v in va_offset]))
def get_split_reads(sample_alignment):
"""Isolate split reads from a sample alignment.
This uses a python script supplied with Lumppy, that is run as a
separate process.
NOTE THAT THIS SCRIPT ONLY WORKS WITH BWA MEM.
"""
bwa_split_dataset = get_dataset_with_type(
sample_alignment, Dataset.TYPE.BWA_SPLIT)
if bwa_split_dataset is not None:
if (bwa_split_dataset.status == Dataset.STATUS.READY and
os.path.exists(bwa_split_dataset.get_absolute_location())):
return bwa_split_dataset
else:
bwa_split_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_SPLIT,
type=Dataset.TYPE.BWA_SPLIT,
status=Dataset.STATUS.NOT_STARTED)
sample_alignment.dataset_set.add(bwa_split_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_split_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_split_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment, Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(bam_filename), "BAM file '%s' is missing." % (
bam_filename)
# NOTE: This assumes the index just adds at .bai, w/ same path otherwise
# - will this always be true?
if not os.path.exists(bam_filename+'.bai'):
index_bam_file(bam_filename)
bam_split_filename = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_split_reads.bam')
# Use lumpy bwa-mem split read script to pull out split reads.
filter_split_reads = ' | '.join([
'{samtools} view -h {bam_filename}',
'python {lumpy_bwa_mem_sr_script} -i stdin',
'{samtools} view -Sb -']).format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=bam_filename,
lumpy_bwa_mem_sr_script=
settings.LUMPY_EXTRACT_SPLIT_READS_BWA_MEM)
try:
bwa_split_dataset.status = Dataset.STATUS.COMPUTING
bwa_split_dataset.save(update_fields=['status'])
with open(bam_split_filename, 'w') as fh:
subprocess.check_call(filter_split_reads,
stdout=fh,
shell=True,
executable=settings.BASH_PATH)
# sort the split reads, overwrite the old file
subprocess.check_call([settings.SAMTOOLS_BINARY, 'sort', bam_split_filename,
os.path.splitext(bam_split_filename)[0]])
_filter_out_interchromosome_reads(bam_split_filename)
bwa_split_dataset.status = Dataset.STATUS.READY
bwa_split_dataset.filesystem_location = clean_filesystem_location(
bam_split_filename)
except subprocess.CalledProcessError:
# if there are no split reads, then fail.
bwa_split_dataset.filesystem_location = ''
bwa_split_dataset.status = Dataset.STATUS.FAILED
bwa_split_dataset.save()
return bwa_split_dataset
def get_discordant_read_pairs(sample_alignment):
"""Isolate discordant pairs of reads from a sample alignment.
"""
# First, check if completed dataset already exists.
bwa_disc_dataset = get_dataset_with_type(
sample_alignment, Dataset.TYPE.BWA_DISCORDANT)
if bwa_disc_dataset is not None:
if (bwa_disc_dataset.status == Dataset.STATUS.READY and
os.path.exists(bwa_disc_dataset.get_absolute_location())):
return bwa_disc_dataset
else:
bwa_disc_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_DISCORDANT,
type=Dataset.TYPE.BWA_DISCORDANT)
sample_alignment.dataset_set.add(bwa_disc_dataset)
# If here, we are going to run or re-run the Dataset.
bwa_disc_dataset.status = Dataset.STATUS.NOT_STARTED
bwa_disc_dataset.save(update_fields=['status'])
bam_dataset = get_dataset_with_type(sample_alignment, Dataset.TYPE.BWA_ALIGN)
bam_filename = bam_dataset.get_absolute_location()
assert os.path.exists(bam_filename), "BAM file '%s' is missing." % (
bam_filename)
# NOTE: This assumes the index just adds at .bai, w/ same path otherwise
# - will this always be true?
if not os.path.exists(bam_filename+'.bai'):
index_bam_file(bam_filename)
bam_discordant_filename = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_discordant_pairs.bam')
try:
bwa_disc_dataset.status = Dataset.STATUS.COMPUTING
bwa_disc_dataset.save(update_fields=['status'])
# Use bam read alignment flags to pull out discordant pairs only
filter_discordant = ' | '.join([
'{samtools} view -u -F 0x0002 {bam_filename} ',
'{samtools} view -u -F 0x0100 - ',
'{samtools} view -u -F 0x0004 - ',
'{samtools} view -u -F 0x0008 - ',
'{samtools} view -b -F 0x0400 - ']).format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=bam_filename)
with open(bam_discordant_filename, 'w') as fh:
subprocess.check_call(filter_discordant,
stdout=fh, shell=True, executable=settings.BASH_PATH)
# sort the discordant reads, overwrite the old file
subprocess.check_call([settings.SAMTOOLS_BINARY, 'sort', bam_discordant_filename,
os.path.splitext(bam_discordant_filename)[0]])
_filter_out_interchromosome_reads(bam_discordant_filename)
bwa_disc_dataset.filesystem_location = clean_filesystem_location(
bam_discordant_filename)
bwa_disc_dataset.status = Dataset.STATUS.READY
except subprocess.CalledProcessError:
bwa_disc_dataset.filesystem_location = ''
bwa_disc_dataset.status = Dataset.STATUS.FAILED
bwa_disc_dataset.save()
return bwa_disc_dataset
def align_with_bwa_mem(alignment_group, sample_alignment):
"""
REPLACES OLD BWA PIPELINE USING ALN AND SAMPE/SAMSE
Aligns a sample to a reference genome using the bwa tool.
Args:
alignment_group: AlignmentGroup that this alignment is part of.
sample_alignment: ExperimentSampleToAlignment. The respective dataset
is assumed to have been created as well.
"""
# Start by gettng fresh objects from database.
sample_alignment = ExperimentSampleToAlignment.objects.get(
id=sample_alignment.id)
experiment_sample = sample_alignment.experiment_sample
alignment_group = AlignmentGroup.objects.get(id=alignment_group.id)
# Grab the reference genome fasta for the alignment.
ref_genome_fasta = get_dataset_with_type(
alignment_group.reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
# Get the BWA Dataset and set it to computing.
bwa_dataset = sample_alignment.dataset_set.get(
type=Dataset.TYPE.BWA_ALIGN)
bwa_dataset.status = Dataset.STATUS.COMPUTING
bwa_dataset.save(update_fields=['status'])
# Create a file that we'll write stderr to.
error_path = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_align.error')
error_output = open(error_path, 'w')
# The alignment group is now officially ALIGNING.
if alignment_group.status != AlignmentGroup.STATUS.ALIGNING:
alignment_group.status = AlignmentGroup.STATUS.ALIGNING
alignment_group.start_time = datetime.now()
alignment_group.end_time = None
alignment_group.save(update_fields=['status', 'start_time', 'end_time'])
error_output.write(
"==START OF ALIGNMENT PIPELINE FOR %s, (%s) ==\n" % (
sample_alignment.experiment_sample.label,
sample_alignment.uid))
# We wrap the alignment logic in a try-except so that if an error occurs,
# we record it and update the status of the Dataset to FAILED if anything
# should fail.
try:
# Build index if the index doesn't exist.
# NOTE: When aligning multiple samples to the same reference genome
# concurrently, the build index method should be called once to completion
# before starting the concurrent alignment jobs.
ensure_bwa_index(ref_genome_fasta)
# Grab the fastq sources, and determine whether we are doing paired ends.
# First, grab fastq1, which must exist
fq1_queryset = experiment_sample.dataset_set.filter(
type=Dataset.TYPE.FASTQ1)
assert fq1_queryset, "Must have at least one .fastq file"
fq1_dataset = fq1_queryset[0]
input_reads_1_fq = fq1_dataset.wrap_if_compressed()
input_reads_1_fq_path = fq1_dataset.get_absolute_location()
# Second, check if fastq2 exists and set is_paired_end
fq2_queryset = experiment_sample.dataset_set.filter(
type=Dataset.TYPE.FASTQ2)
if fq2_queryset:
is_paired_end = True
fq2_dataset = fq2_queryset[0]
input_reads_2_fq = fq2_dataset.wrap_if_compressed()
input_reads_2_fq_path = fq2_dataset.get_absolute_location()
else:
is_paired_end = False
# 1. Generate SA coordinates.
read_fq_1_path, read_fq_1_fn = os.path.split(input_reads_1_fq_path)
align_input_args = ' '.join([
'%s/bwa/bwa' % settings.TOOLS_DIR,
'mem',
'-t', '1', # threads
'-R', '"'+read_group_string(experiment_sample)+'"',
# uncomment this to keep secondary alignments (for finding and marking paralogy regions)
# But before we can uncomment we need to fix de novo assembly code
'-a',
ref_genome_fasta,
input_reads_1_fq,
])
if is_paired_end:
read_fq_2_path, read_fq_2_fn = os.path.split(input_reads_2_fq_path)
align_input_args += ' ' + input_reads_2_fq
# To skip saving the SAM file to disk directly, pipe output directly to
# make a BAM file.
align_input_args += ' | ' + settings.SAMTOOLS_BINARY + ' view -bS -'
### 2. Generate SAM output.
output_bam = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_align.bam')
error_output.write(align_input_args)
# Flush the output here so it gets written before the alignments.
error_output.flush()
with open(output_bam, 'w') as fh:
subprocess.check_call(align_input_args,
stdout=fh, stderr=error_output,
shell=True, executable=settings.BASH_PATH)
# Set processing mask to not compute insert metrics if reads are
# not paired end, as the lumpy script only works on paired end reads
opt_processing_mask = {}
if not is_paired_end:
opt_processing_mask['compute_insert_metrics'] = False
# Do several layers of processing on top of the initial alignment.
result_bam_file = process_sam_bam_file(sample_alignment,
alignment_group.reference_genome, output_bam, error_output,
opt_processing_mask=opt_processing_mask)
# Add the resulting file to the dataset.
bwa_dataset.filesystem_location = clean_filesystem_location(
result_bam_file)
bwa_dataset.save()
# Isolate split and discordant reads for SV calling.
get_discordant_read_pairs(sample_alignment)
get_split_reads(sample_alignment)
# Add track to JBrowse.
add_bam_file_track(alignment_group.reference_genome, sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bwa_dataset.status = Dataset.STATUS.READY
bwa_dataset.save()
delete_redundant_files(sample_alignment.get_model_data_dir())
except:
import traceback
error_output.write(traceback.format_exc())
bwa_dataset.status = Dataset.STATUS.FAILED
bwa_dataset.save()
return
finally:
print error_path
error_output.write('==END OF ALIGNMENT PIPELINE==\n')
error_output.close()
# Add the error Dataset to the object.
error_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_ALIGN_ERROR,
type=Dataset.TYPE.BWA_ALIGN_ERROR,
filesystem_location=clean_filesystem_location(error_path))
sample_alignment.dataset_set.add(error_dataset)
sample_alignment.save()
return sample_alignment
def generate_new_reference_genome(variant_set, new_ref_genome_params):
"""Uses reference_genome_maker code to create a new ReferenceGenome
from the given VariantSet (applies Variants to existing ReferenceGenome.)
Args:
variant_set: The VariantSet from which we'll generate the new
ReferenceGenome.
new_ref_genome_params: Dictionary of params, including:
* label (required): Label for the new ReferenceGenome.
Returns:
The new ReferenceGenome.
Raises:
ValidationException if we don't support this use case.
"""
try:
# Validate / parse params.
assert 'label' in new_ref_genome_params
new_ref_genome_label = new_ref_genome_params['label']
original_ref_genome = variant_set.reference_genome
# Create a ReferenceGenome to track the position.
reference_genome = ReferenceGenome.objects.create(
project=original_ref_genome.project, label=new_ref_genome_label)
# Location for the generated Genbank.
filename_prefix = generate_safe_filename_prefix_from_label(
new_ref_genome_label)
output_root = os.path.join(reference_genome.get_model_data_dir(),
filename_prefix)
full_output_path = clean_filesystem_location(output_root + '.genbank')
# Dataset to track the location.
dataset = Dataset.objects.create(
label=Dataset.TYPE.REFERENCE_GENOME_GENBANK,
type=Dataset.TYPE.REFERENCE_GENOME_GENBANK,
filesystem_location=full_output_path,
status=Dataset.STATUS.NOT_STARTED)
reference_genome.dataset_set.add(dataset)
# Prepare params for calling referece_genome_maker.
# If the old genome is annotated, use it, otherwise, use the FASTA.
# The BioPython SeqRecord should be the same either way.
if original_ref_genome.is_annotated():
original_genome_path = original_ref_genome.dataset_set.get(
type=Dataset.TYPE.REFERENCE_GENOME_GENBANK).\
get_absolute_location()
sequence_record = SeqIO.read(original_genome_path, 'genbank')
else:
original_genome_path = original_ref_genome.dataset_set.get(
type=Dataset.TYPE.REFERENCE_GENOME_FASTA).\
get_absolute_location()
sequence_record = SeqIO.read(original_genome_path, 'fasta')
filename_prefix = generate_safe_filename_prefix_from_label(
new_ref_genome_label)
output_root = os.path.join(reference_genome.get_model_data_dir(),
filename_prefix)
# Create a fake, empty vcf path for now, as we're just getting
# end-to-end to work.
if not os.path.exists(settings.TEMP_FILE_ROOT):
os.mkdir(settings.TEMP_FILE_ROOT)
_, vcf_path = tempfile.mkstemp(suffix='_' + filename_prefix + '.vcf',
dir=settings.TEMP_FILE_ROOT)
with open(vcf_path, 'w') as vcf_fh:
export_variant_set_as_vcf(variant_set, vcf_fh)
dataset.status = Dataset.STATUS.COMPUTING
dataset.save(update_fields=['status'])
try:
new_ref_genome_seq_record = reference_genome_maker.run(
sequence_record, output_root, vcf_path)
except Exception as e:
dataset.status = Dataset.STATUS.FAILED
dataset.save(update_fields=['status'])
raise e
reference_genome.save()
dataset.status = Dataset.STATUS.READY
dataset.save(update_fields=['status'])
# Since the post_add_seq_to_ref_genome() signal couldn't run before,
# we need to make sure to run it now.
prepare_ref_genome_related_datasets(reference_genome, dataset)
return reference_genome
except Exception as e:
raise ValidationException(str(e))
def generate_new_reference_genome(variant_set, new_ref_genome_params):
"""Uses reference_genome_maker code to create a new ReferenceGenome
from the given VariantSet (applies Variants to existing ReferenceGenome.)
Args:
variant_set: The VariantSet from which we'll generate the new
ReferenceGenome.
new_ref_genome_params: Dictionary of params, including:
* label (required): Label for the new ReferenceGenome.
Returns:
The new ReferenceGenome.
Raises:
ValidationException if we don't support this use case.
"""
try:
# Validate / parse params.
assert 'label' in new_ref_genome_params
new_ref_genome_label = new_ref_genome_params['label']
original_ref_genome = variant_set.reference_genome
# Create a ReferenceGenome to track the position.
reference_genome = ReferenceGenome.objects.create(
project=original_ref_genome.project,
label=new_ref_genome_label)
# Location for the generated Genbank.
filename_prefix = generate_safe_filename_prefix_from_label(
new_ref_genome_label)
output_root = os.path.join(reference_genome.get_model_data_dir(),
filename_prefix)
full_output_path = clean_filesystem_location(output_root + '.genbank')
# Dataset to track the location.
dataset = Dataset.objects.create(
label=Dataset.TYPE.REFERENCE_GENOME_GENBANK,
type=Dataset.TYPE.REFERENCE_GENOME_GENBANK,
filesystem_location=full_output_path,
status=Dataset.STATUS.NOT_STARTED)
reference_genome.dataset_set.add(dataset)
# Prepare params for calling referece_genome_maker.
original_fasta_path = original_ref_genome.dataset_set.get(
type=Dataset.TYPE.REFERENCE_GENOME_FASTA).\
get_absolute_location()
sequence_record = SeqIO.read(original_fasta_path, 'fasta')
filename_prefix = generate_safe_filename_prefix_from_label(
new_ref_genome_label)
output_root = os.path.join(reference_genome.get_model_data_dir(),
filename_prefix)
# Create a fake, empty vcf path for now, as we're just getting
# end-to-end to work.
if not os.path.exists(settings.TEMP_FILE_ROOT):
os.mkdir(settings.TEMP_FILE_ROOT)
_, vcf_path = tempfile.mkstemp(
suffix='_' + filename_prefix + '.vcf',
dir=settings.TEMP_FILE_ROOT)
with open(vcf_path, 'w') as vcf_fh:
export_variant_set_as_vcf(variant_set, vcf_fh)
dataset.status = Dataset.STATUS.COMPUTING
dataset.save(update_fields=['status'])
try:
new_ref_genome_seq_record = reference_genome_maker.run(
sequence_record, output_root, vcf_path)
except Exception as e:
dataset.status = Dataset.STATUS.FAILED
dataset.save(update_fields=['status'])
raise e
reference_genome.save()
dataset.status = Dataset.STATUS.READY
dataset.save(update_fields=['status'])
# Since the post_add_seq_to_ref_genome() signal couldn't run before,
# we need to make sure to run it now.
prepare_ref_genome_related_datasets(reference_genome, dataset)
return reference_genome
except Exception as e:
raise ValidationException(str(e))
def derivation_fn(sample_alignment, new_dataset):
"""Creates a bam file with reads relevant for de novo assembly.
"""
# Get the original bam file.
bam_dataset = get_dataset_with_type(sample_alignment,
Dataset.TYPE.BWA_ALIGN)
orig_bam_filename = bam_dataset.get_absolute_location()
# Allocate a filename for the new dataset.
de_novo_bam_filelocation = (os.path.splitext(orig_bam_filename)[0] +
'.de_novo.bam')
new_dataset.filesystem_location = clean_filesystem_location(
de_novo_bam_filelocation)
new_dataset.save(update_fields=['filesystem_location'])
### Strategy
# 0. Create intermediate sam
# 1. Grab unmapped reads.
# a. If the corresponding pair for any read was mapped (and thus not
# in the unmapped dataset), grab that from the original bam file.
# 2. Grab split reads.
# 2b. Add reads that are in intervals of interest.
# 3. Sort by name so that paired reads are next to each other
# 4. Filter out duplicate reads. Requires sort in step 3.
# 5. Convert to bam.
# 6. Delete intermediate files.
# 0. Create intermediate files.
intermediate_sam = (
os.path.splitext(de_novo_bam_filelocation)[0] + '.int.sam')
paired_intermediate_sam = (
os.path.splitext(intermediate_sam)[0] + '.paired.sam')
sorted_intermediate_sam = (
os.path.splitext(paired_intermediate_sam)[0] + '.sorted.sam')
deduped_sorted_intermediate_sam = (
os.path.splitext(sorted_intermediate_sam)[0] + '.deduped.sam')
intermediate_files = [
intermediate_sam,
paired_intermediate_sam,
sorted_intermediate_sam,
deduped_sorted_intermediate_sam
]
# 1. Get unmapped reads.
unmapped_reads_dataset = get_unmapped_reads(sample_alignment)
unmapped_reads_bam_file = unmapped_reads_dataset.get_absolute_location()
cmd = '{samtools} view -h {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=unmapped_reads_bam_file)
with open(intermediate_sam, 'w') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
# 2. Grab split reads.
split_reads_dataset = get_split_reads(sample_alignment)
split_rads_bam_file = split_reads_dataset.get_absolute_location()
cmd = '{samtools} view {bam_filename}'.format(
samtools=settings.SAMTOOLS_BINARY,
bam_filename=split_rads_bam_file)
with open(intermediate_sam, 'a') as output_fh:
subprocess.check_call(cmd, stdout=output_fh, shell=True)
# 2b. Add reads that are in any of intervals that we want to include.
if force_include_reads_in_intervals:
with open(intermediate_sam, 'a') as output_fh:
get_reads_in_interval_list(orig_bam_filename,
force_include_reads_in_intervals, output_fh)
# 2c. For each of the reads that we've included, grab their
# corresponding pairs.
add_paired_mates(intermediate_sam, orig_bam_filename,
paired_intermediate_sam)
# 3. Sort by name so that paired reads are next to each other
cmd = (
'(grep ^"@" {sam_file}; grep -v ^"@" {sam_file} | '
'sort -k1,1 -k2,2n) > {sorted_sam_file}'
).format(
sam_file=paired_intermediate_sam,
sorted_sam_file=sorted_intermediate_sam)
subprocess.call(cmd, shell=True)
# 4. Filter out duplicate reads. Requires sort in step 3.
cmd = 'uniq {sorted_sam_file} > {deduped_sam_file}'.format(
sorted_sam_file=sorted_intermediate_sam,
deduped_sam_file=deduped_sorted_intermediate_sam)
subprocess.call(cmd, shell=True)
# 5. Convert to bam.
cmd = '{samtools} view -Sb {sam_file} > {final_bam_file}'.format(
samtools=settings.SAMTOOLS_BINARY,
sam_file=intermediate_sam,
final_bam_file=de_novo_bam_filelocation)
subprocess.call(cmd, shell=True)
# 6. Delete intermediate files.
for f in intermediate_files:
os.remove(f)
def align_with_bwa_mem(alignment_group, sample_alignment):
"""
REPLACES OLD BWA PIPELINE USING ALN AND SAMPE/SAMSE
Aligns a sample to a reference genome using the bwa tool.
Args:
alignment_group: AlignmentGroup that this alignment is part of.
sample_alignment: ExperimentSampleToAlignment. The respective dataset
is assumed to have been created as well.
"""
# Start by gettng fresh objects from database.
sample_alignment = ExperimentSampleToAlignment.objects.get(
id=sample_alignment.id)
experiment_sample = sample_alignment.experiment_sample
alignment_group = AlignmentGroup.objects.get(id=alignment_group.id)
# Grab the reference genome fasta for the alignment.
ref_genome_fasta = get_dataset_with_type(
alignment_group.reference_genome,
Dataset.TYPE.REFERENCE_GENOME_FASTA).get_absolute_location()
# Get the BWA Dataset and set it to computing.
bwa_dataset = sample_alignment.dataset_set.get(type=Dataset.TYPE.BWA_ALIGN)
bwa_dataset.status = Dataset.STATUS.COMPUTING
bwa_dataset.save(update_fields=['status'])
# Create a file that we'll write stderr to.
error_path = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_align.error')
error_output = open(error_path, 'w')
# The alignment group is now officially ALIGNING.
if alignment_group.status != AlignmentGroup.STATUS.ALIGNING:
alignment_group.status = AlignmentGroup.STATUS.ALIGNING
alignment_group.start_time = datetime.now()
alignment_group.end_time = None
alignment_group.save(
update_fields=['status', 'start_time', 'end_time'])
error_output.write(
"==START OF ALIGNMENT PIPELINE FOR %s, (%s) ==\n" %
(sample_alignment.experiment_sample.label, sample_alignment.uid))
# We wrap the alignment logic in a try-except so that if an error occurs,
# we record it and update the status of the Dataset to FAILED if anything
# should fail.
try:
# Build index if the index doesn't exist.
# NOTE: When aligning multiple samples to the same reference genome
# concurrently, the build index method should be called once to completion
# before starting the concurrent alignment jobs.
ensure_bwa_index(ref_genome_fasta)
# Grab the fastq sources, and determine whether we are doing paired ends.
# First, grab fastq1, which must exist
fq1_queryset = experiment_sample.dataset_set.filter(
type=Dataset.TYPE.FASTQ1)
assert fq1_queryset, "Must have at least one .fastq file"
fq1_dataset = fq1_queryset[0]
input_reads_1_fq = fq1_dataset.wrap_if_compressed()
input_reads_1_fq_path = fq1_dataset.get_absolute_location()
# Second, check if fastq2 exists and set is_paired_end
fq2_queryset = experiment_sample.dataset_set.filter(
type=Dataset.TYPE.FASTQ2)
if fq2_queryset:
is_paired_end = True
fq2_dataset = fq2_queryset[0]
input_reads_2_fq = fq2_dataset.wrap_if_compressed()
input_reads_2_fq_path = fq2_dataset.get_absolute_location()
else:
is_paired_end = False
# 1. Generate SA coordinates.
read_fq_1_path, read_fq_1_fn = os.path.split(input_reads_1_fq_path)
align_input_args = ' '.join([
'%s/bwa/bwa' % settings.TOOLS_DIR,
'mem',
'-t',
'1', # threads
'-R',
'"' + read_group_string(experiment_sample) + '"',
# uncomment this to keep secondary alignments (for finding and marking paralogy regions)
# But before we can uncomment we need to fix de novo assembly code
'-a',
ref_genome_fasta,
input_reads_1_fq,
])
if is_paired_end:
read_fq_2_path, read_fq_2_fn = os.path.split(input_reads_2_fq_path)
align_input_args += ' ' + input_reads_2_fq
# To skip saving the SAM file to disk directly, pipe output directly to
# make a BAM file.
align_input_args += ' | ' + settings.SAMTOOLS_BINARY + ' view -bS -'
### 2. Generate SAM output.
output_bam = os.path.join(sample_alignment.get_model_data_dir(),
'bwa_align.bam')
error_output.write(align_input_args)
# Flush the output here so it gets written before the alignments.
error_output.flush()
with open(output_bam, 'w') as fh:
subprocess.check_call(align_input_args,
stdout=fh,
stderr=error_output,
shell=True,
executable=settings.BASH_PATH)
# Set processing mask to not compute insert metrics if reads are
# not paired end, as the lumpy script only works on paired end reads
opt_processing_mask = {}
if not is_paired_end:
opt_processing_mask['compute_insert_metrics'] = False
# Do several layers of processing on top of the initial alignment.
result_bam_file = process_sam_bam_file(
sample_alignment,
alignment_group.reference_genome,
output_bam,
error_output,
opt_processing_mask=opt_processing_mask)
# Add the resulting file to the dataset.
bwa_dataset.filesystem_location = clean_filesystem_location(
result_bam_file)
bwa_dataset.save()
# Isolate split and discordant reads for SV calling.
get_discordant_read_pairs(sample_alignment)
get_split_reads(sample_alignment)
# Add track to JBrowse.
add_bam_file_track(alignment_group.reference_genome, sample_alignment,
Dataset.TYPE.BWA_ALIGN)
bwa_dataset.status = Dataset.STATUS.READY
bwa_dataset.save()
delete_redundant_files(sample_alignment.get_model_data_dir())
except:
import traceback
error_output.write(traceback.format_exc())
bwa_dataset.status = Dataset.STATUS.FAILED
bwa_dataset.save()
return
finally:
print error_path
error_output.write('==END OF ALIGNMENT PIPELINE==\n')
error_output.close()
# Add the error Dataset to the object.
error_dataset = Dataset.objects.create(
label=Dataset.TYPE.BWA_ALIGN_ERROR,
type=Dataset.TYPE.BWA_ALIGN_ERROR,
filesystem_location=clean_filesystem_location(error_path))
sample_alignment.dataset_set.add(error_dataset)
sample_alignment.save()
return sample_alignment
